（五）水稻遗传研究动态（牵头与起草专家：庄杰云博士等）
1、近 5 年水稻遗传研究概述
2、2007 年我国水稻遗传研究动态研究
3、2007 年国际水稻遗传研究动态
4、水稻遗传研究发展趋势
001《Plant Cell Reports》
The rice pathogen-related protein 10 (JIOsPR10) is induced by abiotic and biotic stresses and exhibits ribonuclease activity
Sun Tae Kim, Seok Yu, Young Hyun Kang, Sang Gon Kim, Jae-Yean Kim, Sun-Hyung Kim, Kyu Young Kang Environmental Biotechnology
National Core Research Center, Gyeongsang National University, Jinju, 660-701, Korea
Abstract:We previously reported that rice blast fungus or jasmonic acid induced the expression of rice pathogenesis-related class 10 (JIOsPR10)
proteins (Kim et al. 2003, 2004). However, no further studies have been carried out to examine the expression, localization, and enzymatic activity
of this protein in either developmental tissues or in tissues under abiotic stress conditions. In this study, rice JIOsPR10 was examined by Western
blot analysis, immunolocalization, and biochemical assays. Western blots revealed that the JIOsPR10 protein was expressed in developmental
tissues, including in flower and root. The protein was also expressed under abiotic stresses, such as occurs during senescence and wounding. Using
immunohistochemical techniques, we determined that expression of JIOsPR10 was localized to the palea of flower, in the exodermis, and inner
part of the endodermis of the root. In senescencing tissues of leaf and coleoptiles, its expression was localized in vascular bundles. The RNase
activity using JIOsPR10 recombinant protein was determined and abolished after treatment with DTT in a native in-gel assay. To test this, we
created JIOsPR10 mutant proteins containing serine substitutions of amino acids C81S, C83S, or both and examined their RNase activities. The
activity of the C83S mutant was decreased in the agarose gel assay compared to the wild type. Taken together, we hypothesize that the JIOsPR10
protein possesses RNase activity that is sensitive to DTT, suggesting the importance of the disulfide bonding between cysteine residues and that it
might play a role in constitutive self-defense mechanisms in plants against biotic and abiotic stresses.
002《Planta》
Expression of rice heat stress transcription factor OsHsfA2e enhances tolerance to environmental stresses in transgenic Arabidopsis
Naoki Yokotani, Takanari Ichikawa, Youichi Kondou, Minami Matsui, Hirohiko Hirochika, Masaki Iwabuchi, Kenji Oda
Research Institute for Biological Sciences (RIBS), Okayama, 7549-1 Kibichuo-cho, Kaga-gun, Okayama 716-1241, Japan
Abstract:Plant growth and crop yields are limited by high-temperature stresses. In this study, we attempted to isolate the rice genes responsible for
high-temperature stress tolerance using a transformed Arabidopsis population expressing a full-length cDNA library of rice. From approximately
20,000 lines of transgenic Arabidopsis, we isolated a thermotolerant line, R04333, that could survive transient heat stress at the cotyledon stage.
The rice cDNA inserted in R04333 encodes OsHsfA2e, a member of the heat stress transcription factors. The thermotolerant phenotype was
observed in newly constructed transgenic Arabidopsis plants expressing OsHsfA2e. Among 5 A2-type HSF genes encoded in the rice genome, four
genes, including OsHsfA2e, are induced by high temperatures in rice seedlings. The OsHsfA2e protein was localized to the nuclear region and
exhibited transcription activation activity in the C-terminal region. Microarray analysis demonstrated that under unstressed conditions transgenic
Arabidopsis overexpressing OsHsfA2e highly expressed certain stress-associated genes, including several classes of heat-shock proteins. The
thermotolerant phenotype was observed not only in the cotyledons but also in rosette leaves, inflorescence stems and seeds. In addition, transgenic
Arabidopsis exhibited tolerance to high-salinity stress. These observations suggest that the OsHsfA2e may be useful in molecular breeding
designed to improve the environmental stress tolerance of crops.
003《Journal of General Plant Pathology》
Suppression of rice blast using freeze-killed mycelia of biocontrol fungal candidate MKP5111B
Nobuaki Ohtaka, Hitoshi Kawamata, Kazuhiko Narisawa
Plant Biotechnology Institute, Ibaraki Agricultural Center, Ago, Kasama, Ibaraki 319-0292, Japan
Abstract:Several applications of the suppressive fungus MKP5111B, isolated from the phylloplane of rice plants, were tested in an effort to control
rice blast disease. Three treatments with MKP5111B [living (Std), killed with liquid nitrogen (FR), and autoclaved (AC)] were either sprayed onto
rice seedlings or mixed into seed-sown soil. Three weeks after spraying and 4 weeks after the soil application, we introduced Magnaporthe oryzae,
the causal agent of rice blast, into our systems. The Std and FR treatments suppressed rice blast, but the AC treatment proved ineffective. Although
a suppressive effect was seen on new leaves, no mycelium of MKP5111B was seen. The fungus thus may have induced a systemic resistance in the
rice plants. A substance from MKP5111B, such as elicitor molecule(s) are likely responsible for the induced resistance.
004《Plant Molecular Biology》
Identification and localisation of the rice nicotianamine aminotransferase gene OsNAAT1 expression suggests the site of phytosiderophore
synthesis in rice
Haruhiko Inoue, Michiko Takahashi, Takanori Kobayashi, Motofumi Suzuki, Hiromi Nakanishi, Satoshi Mori, Naoko K. Nishizawa
Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
Abstract:Rice plants (Oryza sativa L.) take up iron using iron-chelating compounds known as mugineic acid family phytosiderophores (MAs). In
the biosynthetic pathway of MAs, nicotianamine aminotransferase (NAAT) catalyses the key step from nicotianamine to the 3''-keto form. In the
present study, we identified six rice NAAT genes (OsNAAT1-6) by screening a cDNA library made from Fe-deficient rice roots and by searching
databases. Among the NAAT homologues, OsNAAT1 belongs to a subgroup containing barley functional NAAT (HvNAAT-A and HvNAAT-B) as
well as a maize homologue cloned by cDNA library screening (ZmNAAT1). Northern blot and RT-PCR analysis showed that OsNAAT1, but not
OsNAAT2-6, was strongly up-regulated by Fe deficiency, both in roots and shoots. The OsNAAT1 protein had NAAT enzyme activity in vitro,
confirming that the OsNAAT1 gene encodes functional NAAT. Promoter-GUS analysis revealed that OsNAAT1 was expressed in companion and
pericycle cells adjacent to the protoxylem of Fe-sufficient roots. In addition, expression was induced in all cells of Fe-deficient roots, with
particularly strong GUS activity evident in the companion and pericycle cells. OsNAAT1 expression was also observed in the companion cells of
Fe-sufficient shoots, and was clearly induced in all the cells of Fe-deficient leaves. These expression patterns highly resemble those of OsNAS1,
OsNAS2 and OsDMAS1, the genes responsible for MAs biosynthesis for Fe acquisition. These findings strongly suggest that rice synthesises MAs
in whole Fe-deficient roots to acquire Fe from the rhizosphere, and also in phloem cells to maintain metal homeostasis facilitated by
MAs-mediated long-distance transport.
005《Biologia Plantarum》
Root morphology and cadmium uptake kinetics of the cadmium-sensitive rice mutant
J. Y. He, C. Zhu, Y. F. Ren, D. A. Jiang, Z. X. Sun
College of Life Science, Zhejiang University, Hangzhou, 310058, P.R. China
Abstract:To understand the physiological mechanism that confers Cd sensitivity, root morphology and Cd uptake kinetics of the Cd-sensitive
mutant and wild type rice were investigated. The root length, root surface area, and root number of mutant rice decreased more significantly with
increasing Cd concentration in growth media compared with the wild type rice. The uptake kinetics for 109Cd2+ in roots of both the mutant and wild
type rice were characterized by a rapid linear phase during the first 6 h and a slower linear phase during the subsequent period.
Concentration-dependent Cd2+ influx in both species could be characterized by the Michaelis-Menten equation, with similar apparent Km values
for mutant and wild type rice (2.54 and 2.37 µM, respectively). However, the V max for Cd2+ influx in mutant root cells was nearly 2-fold higher
than that for wild type rice, indicating that enhanced absorption into the root is one of the mechanisms involved in Cd sensitivity in mutant rice.
006《Annals of Botany》
Progress in Research and Development on Hybrid Rice: A Super-domesticate in China
Shi-Hua Cheng, Jie-Yun Zhuang, Ye-Yang Fan, Jing-Hong Du, and Li-Yong Cao
Chinese National Center for Rice Improvement and State Key Laboratory of Rice Biology, China National Rice Research Institute (CNRRI),
Hangzhou 310006, China
Abstract:
Background: China has been successful in breeding hybrid rice strains, but is now facing challenges to develop new hybrids with high-yielding
potential, better grain quality, and tolerance to biotic and abiotic stresses. This paper reviews the most significant advances in hybrid rice breeding
in China, and presents a recent study on fine-mapping quantitative trait loci (QTLs) for yield traits.
Scope: By exploiting new types of male sterility, hybrid rice production in China has become more diversified. The use of inter-subspecies crosses
has made an additional contribution to broadening the genetic diversity of hybrid rice and played an important role in the breeding of super rice
hybrids in China. With the development and application of indica-inclined and japonica-inclined parental lines, new rice hybrids with super
high-yielding potential have been developed and are being grown on a large scale. DNA markers for subspecies differentiation have been identified
and applied, and marker-assisted selection performed for the development of restorer lines carrying disease resistance genes. The genetic basis of
heterosis in highly heterotic hybrids has been studied, but data from these studies are insufficient to draw sound conclusions. In a QTL study using
stepwise residual heterozygous lines, two linked intervals harbouring QTLs for yield traits were resolved, one of which was delimited to a 125-kb
region.
Conclusions: Advances in rice genomic research have shed new light on the genetic study and germplasm utilization in rice. Molecular
marker-assisted selection is a powerful tool to increase breeding efficiency, but much work remains to be done before this technique can be
extended from major genes to QTLs
007《Applied Microbiology and Biotechnology》
Biologically active components and nutraceuticals in the Monascus -fermented rice: a review
Yii-Lih Lin, Teng-Hsu Wang, Min-Hsiung Lee, Nan-Wei Su
Department of Agricultural Chemistry, National Taiwan University, no. 1, Sec. 4, Roosevelt Rd., Taipei, 10617, Taiwan，China
Abstract:Monascus-fermented rice has traditionally been used as a natural food colorant and food preservative of meat and fish for centuries. It has
recently become a popular dietary supplement because of many of its bioactive constituents being discovered, including a series of active drug
compounds, monacolins, indicated as the 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors for reducing serum cholesterol level. The
controversy of its safety has been provoked because a mycotoxin, citrinin, is also produced along with the Monascus secondary metabolites by
certain strains or under certain cultivation conditions. This review introduces the basic production process and addresses on the compounds with
bioactive functions. Current advances in avoiding the harmful ingredient citrinin are also discussed.
008《Molecular Biology Reports》
Cloning and functional identification of two members of the ZIP (Zrt, Irt-like protein) gene family in rice ( Oryza sativa L.)
Xia Yang, Ji Huang, Yan Jiang, Hong-Sheng Zhang
State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, Nanjing, 210095, China
Abstract:Two ZIP (Zrt, Irt-like Protein) cDNAs were isolated from rice (Oryza sativa L.) by RT-PCR approach, and named as OsZIP7a and
OsZIP8 respectively. The predicted proteins of OsZIP7a and OsZIP8 consist of 384 and 390 amino acid residues respectively, and display high
similarity to other plant ZIP proteins. Each protein contains eight transmembrane (TM) domains and a highly conserved ZIP signature motif, with
a histidine-rich region in the variable region between TM domains III and IV. By semi-quantitative RT-PCR approach, it was found that the
expression of OsZIP7a was significantly induced in rice roots by iron-deficiency, while that of OsZIP8 induced in both rice roots and shoots by
zinc-deficiency. When expressed in yeast cells, OsZIP7a and OsZIP8 could complement an iron-uptake-deficient yeast mutant and a
zinc-uptake-deficient yeast mutant respectively. It suggested that the OsZIP7a and OsZIP8 might encode an iron and a zinc transporter protein in
rice respectively.
009《Plant Molecular Biology》
Are Pseudokirchneriella subcapitata and Chlorella vulgaris Affected by Environmental Samples from a Rice Field?
Haruhiko Inoue, Michiko Takahashi, Takanori Kobayashi, Motofumi Suzuki, Hiromi Nakanishi, Satoshi Mori, Naoko K. Nishizawa
Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
Abstract:Rice plants (Oryza sativa L.) take up iron using iron-chelating compounds known as mugineic acid family phytosiderophores (MAs). In
the biosynthetic pathway of MAs, nicotianamine aminotransferase (NAAT) catalyses the key step from nicotianamine to the 3''-keto form. In the
present study, we identified six rice NAAT genes (OsNAAT1-6) by screening a cDNA library made from Fe-deficient rice roots and by searching
databases. Among the NAAT homologues, OsNAAT1 belongs to a subgroup containing barley functional NAAT (HvNAAT-A and HvNAAT-B) as
well as a maize homologue cloned by cDNA library screening (ZmNAAT1). Northern blot and RT-PCR analysis showed that OsNAAT1, but not
OsNAAT2-6, was strongly up-regulated by Fe deficiency, both in roots and shoots. The OsNAAT1 protein had NAAT enzyme activity in vitro,
confirming that the OsNAAT1 gene encodes functional NAAT. Promoter-GUS analysis revealed that OsNAAT1 was expressed in companion and
pericycle cells adjacent to the protoxylem of Fe-sufficient roots. In addition, expression was induced in all cells of Fe-deficient roots, with
particularly strong GUS activity evident in the companion and pericycle cells. OsNAAT1 expression was also observed in the companion cells of
Fe-sufficient shoots, and was clearly induced in all the cells of Fe-deficient leaves. These expression patterns highly resemble those of OsNAS1,
OsNAS2 and OsDMAS1, the genes responsible for MAs biosynthesis for Fe acquisition. These findings strongly suggest that rice synthesises MAs
in whole Fe-deficient roots to acquire Fe from the rhizosphere, and also in phloem cells to maintain metal homeostasis facilitated by
MAs-mediated long-distance transport.
010《Euphytica》
Breeding of transgenic rice restorer line for multiple resistance against bacterial blight, striped stem borer and herbicide
Yipei Wei, Fangyin Yao, Changxiang Zhu, Mingsong Jiang, Guangxian Li, Yunzhi Song, Fujiang Wen
State Key Laboratory of Crop Biology, Shandong Agricultural University, Tai'an, 271018, P.R. China
Abstract:In this paper, we described the breeding of transgenic rice restorer line for multiple resistance against bacterial blight, striped stem borer
(SSB) and herbicide by conventional crossing of two transgenic parental lines transformed independently with different genes. Two stable
transgenic rice lines used as donor parents were developed, one was Zhongguo91 which contained cry1Ab gene (for insect resistance) and bar
gene (for tolerance of herbicide), and the other was Yujing6 which contained Xa21 gene (resistance to bacterial blight). The elite restorer line
Hui773 was used as recipient and crossed with the two stable transgenic rice lines. Then five successive backcrosses were made using Hui773 as
recurrent parent. Two rice elite restorers, T773-1 expressing cry1Ab and bar genes and T773-2 expressing Xa21 gene, were obtained, which were
confirmed by PCR analysis and testing selectable marker genes in the hybrid progenies. The cross was made between T773-1 and T773-2 to select
stable restorer line carrying Xa21, cry1Ab and bar genes. Finally, we obtained transgenic restorer line T773 with good agronomic traits and
obvious multiple resistance to Xanthomonas oryzae pv. oryzae, striped stem borer (Chilo suppressalis) and herbicide. The hybrid F1 generation
produced from the cross between transgenic restorer line T773 and a corresponding male sterile line Zaohua2A maintained obvious resistance to
rice bacterial blight, rice leaffolder and striped stem borer, and showed significant heterosis. Our results indicate that it is feasible to develop
transgenic hybrid rice cultivar through breeding transgenic restorer lines.
011《Functional & Integrative Genomics》
Chloroplast DNA insertions into the nuclear genome of rice: the genes, sites and ages of insertion involved
Xingyi Guo, Songlin Ruan, Weiming Hu, Daguang Cai, Longjiang Fan
Institute of Crop Science, Zhejiang University, Hangzhou, 310029, China
Abstract:Rice (Oryza sativa) is one of three predominant grain crops, and its nuclear and organelle genomes have been sequenced. Following
genome analysis revealed many exchanges of DNA sequences between the nuclear and organelle genomes. In this study, a total of 45 chloroplast
DNA insertions more than 2 kb in length were detected in rice nuclear genome. A homologous recombination mechanism is expected for those
chloroplast insertions with high similarity between their flanking sequences. Only five chloroplast insertions with high sequence similarity
between two flanking sequences from an insertion were found in the 45 insertions, suggesting that rice might follow the non-homologous
end-joining (NHEJ) repair of double-stranded breaks mechanism, which is suggested to be common to all eukaryotes. Our studies indicate that the
most chloroplast insertions occurred at a nuclear region characterized by a sharp change of repetitive sequence density. One potential explanation
is that regions such as this might be susceptible target sites or “hotspots” of DNA damage. Our results also suggest that the insertion of
retrotransposon elements or non-chloroplast DNA into chloroplast DNA insertions may contribute significantly to their fragmentation process.
Moreover, based on chloroplast insertions in nuclear genomes of two subspecies (indica and japonica) of cultivated rice, our results strongly
suggest that they diverged during 0.06-0.22 million years ago.
012《Biotechnology Letters》
Construction of a bacterial artificial chromosome (BAC) library of common wild rice ( Oryza rufipogon Griff.) for map-based cloning of genes
selected during the domestication of rice
Xianran Li, LuBin Tan, Haiyan Huang, Zuofeng Zhu, Chenji Li, Songnian Hu, Chuanqing Sun
Department of Plant Genetics and Breeding and State Key Laboratory of Plant Physiology and Biochemistry, China Agricultural University,
Beijing, 100094, China
Abstract:As a prerequisite for the map-based cloning of genes from common wild rice (Oryza rufipogon Griff.), which plays an important role in
the domestication of cultivated rice (O. sativa L.), we constructed a median-insert size bacterial artificial chromosome (BAC) library of the
common wild rice isolate, YJCWR, collected from Yuanjiang, Yunnan Province, China. The library consists of 52,992 clones, with an average
insert size of 50 kb, and all clones were pooled into 46 three-dimensional super-pools to facilitate library screening through the PCR method.
Seventeen candidate clones were isolated by five markers and some clones containing putative target regions were sequenced. Furthermore, in
analyzing the sequences of YJCWR, a retrotransposon, SZ-55, that might contribute to the evolution of Oryza was found.
013《Plant Molecular Biology》
A genome-wide survey of HD-Zip genes in rice and analysis of drought-responsive family members
Adamantia Agalou, Sigit Purwantomo, Elin Övernäs, Henrik Johannesson, Xiaoyi Zhu, Amy Estiati, Rolf J. de Kam, Peter Engström, Inez H.
Slamet-Loedin, Zhen Zhu, Mei Wang, Lizhong Xiong, Annemarie H. Meijer, Pieter B. F. Ouwerkerk
Institute of Biology, Clusius Laboratory, Leiden University, P.O. Box 9505, 2300 RA Leiden, The Netherlands
Abstract:The homeodomain leucine zipper (HD-Zip) genes encode transcription factors that have diverse functions in plant development and have
often been implicated in stress adaptation. The HD-Zip genes are the most abundant group of homeobox (HB) genes in plants and do not occur in
other eukaryotes. This paper describes the complete annotation of the HD-Zip families I, II and III from rice and compares these gene families
with Arabidopsis in a phylogeny reconstruction. Orthologous pairs of rice and Arabidopsis HD-Zip genes were predicted based on neighbour
joining and maximum parsimony (MP) trees with support of conserved intron-exon organization. Additionally, a number of HD-Zip genes
appeared to be unique to rice. Searching of EST and cDNA databases and expression analysis using RT-PCR showed that 30 out of 31 predicted
rice HD-Zip genes are expressed. Most HD-Zip genes were broadly expressed in mature plants and seedlings, but others showed more organ
specific patterns. Like in Arabidopsis and other dicots, a subset of the rice HD-Zip I and II genes was found to be regulated by drought stress. We
identified both drought-induced and drought-repressed HD-Zip genes and demonstrate that these genes are differentially regulated in
drought-sensitive versus drought-tolerant rice cultivars. The drought-repressed HD-Zip family I gene, Oshox4, was selected for promoter-GUS
analysis, showing that drought-responsiveness of Oshox4 is controlled by the promoter and that Oshox4 expression is predominantly
vascular-specific. Loss-of-function analysis of Oshox4 revealed no specific phenotype, but overexpression analysis suggested a role for Oshox4 in
elongation and maturation processes.
014《Chinese Science Bulletin》
Fine mapping and candidate gene analysis of purple pericarp gene Pb in rice ( Oryza sativa L.)
Caixia Wang, Qingyao Shu
IAEA-Zhejiang University Collaborating Center, National Key Laboratory of Rice Biology and Key Laboratory of Chinese Ministry of
Agriculture for Nuclear-Agricultural Sciences, Institute of Nuclear Agricultural Sciences, Zhejiang University, Hangzhou, 310029, China
Abstract:Purple rice is a type of rice with anthocyanins deposited in its grain pericarp. The rice Pb gene controlling purple pericarp character is
known to be on chromosome 4, and the purple color is dominant over white color. In this study, we fine mapped the Pb gene using two F2
segregating populations, i.e. Pei’ai 64S (white) × Yunanheixiannuo (purple) and Pei’ai 64S × Chuanheinuo (purple). In the first-pass mapping, the
Pb gene was located in the region downstream the SSR marker RM3820. In the fine mapping, the candidate region was saturated with InDel and
CAPS markers developed specifically for this study. Eventually, the Pb gene was mapped within the 25-kb region delimited by the upstream
marker RID3 and the downstream marker RID4. The delimited region contained two annotated genes, Ra and bhlh16 (TIGR Rice Genome, R.5).
The former is a homologue of the Myc transcription factor Lc controlling anthocyanin biosynthesis in maize, and the latter is a homologue of the
TT8 gene, which is also an Myc transcription factor gene controlling the pericarp pigmentation in Arabidopsis thaliana. Sequence analysis showed
that the exon 7 of the Ra gene of Yunanheixiannuo and Chuanheinuo had a 2-bp (GT) deletion compared with those of the white rice varieties
Pei’ai 64S, 9311 and Nipponbare. A CAPS marker, CAPSRa, was developed according to the GT deletion for analysis of the two F2 segregating
populations and 106 rice lines. The results showed that all F 2 plants with white pericarp, and all non-purple rice lines (63 white and 22 red)
contained no GT deletion, but all 20 purple rice lines contained the GT deletion. These results suggested that the Ra gene may be the Pb gene and
the purple pericarp characteristic of rice is caused by the GT deletion within exon 7 of the Ra gene.
015《Molecular Genetics and Genomics》
Centromeric retrotransposon lineages predate the maize/rice divergence and differ in abundance and activity
Anupma Sharma, Gernot G. Presting
Department of Molecular Biosciences and Bioengineering, University of Hawaii, 1955 East-West Road, Ag Science Bldg. Rm 218, Honolulu, HI
96822, USA
Abstract:Centromeric retrotransposons (CR) are located almost exclusively at the centromeres of plant chromosomes. Analysis of the emerging
Zea mays inbred B73 genome sequence revealed two novel subfamilies of CR elements of maize (CRM), bringing the total number of known
CRM subfamilies to four. Orthologous subfamilies of each of these CRM subfamilies were discovered in the rice lineage, and the orthologous
relationships were demonstrated with extensive phylogenetic analyses. The much higher number of CRs in maize versus Oryza sativa is due
primarily to the recent expansion of the CRM1 subfamily in maize. At least one incomplete copy of a CRM1 homolog was found in O. sativa ssp.
indica and O. officinalis, but no member of this subfamily could be detected in the finished O. sativa ssp. japonica genome, implying loss of this
prolific subfamily in that subspecies. CRM2 and CRM3, as well as the corresponding rice subfamilies, have been recently active but are present in
low numbers. CRM3 is a full-length element related to the non-autonomous CentA, which is the first described CRM. The oldest subfamily
(CRM4), as well as its rice counterpart, appears to contain only inactive members that are not located in currently active centromeres. The
abundance of active CR elements is correlated with chromosome size in the three plant genomes for which high quality genomic sequence is
available, and the emerging picture of CR elements is one in which different subfamilies are active at different evolutionary times. We propose a
model by which CR elements might influence chromosome and genome size.
016《Theoretical and Applied Genetics》
The QTL analysis on maternal and endosperm genome and their environmental interactions for characters of cooking quality in rice ( Oryza sativa
L.)
X. Zheng, J. G. Wu, X. Y. Lou, H. M. Xu, C. H. Shi
Department of Agronomy, College of Agriculture and Biotechnology, Zhejiang University, 310029 Hangzhou, People's Republic of China
Abstract:Investigations to identify quantitative trait loci (QTLs) governing cooking quality traits including amylose content, gel consistency and
gelatinization temperature (expressed by the alkali spread value) were conducted using a set of 241 RIL populations derived from an elite hybrid
cross of “Zhenshan 97” × “Minghui 63” and their reciprocal backcrosses BC1F1 and BC2F1 populations in two environments. QTLs and QTL ×
environment interactions were analyzed by using the genetic model with endosperm and maternal effects and environmental interaction effects on
quantitative traits of seed in cereal crops. The results suggested that a total of seven QTLs were associated with cooking quality of rice, which were
subsequently mapped to chromosomes 1, 4 and 6. Six of these QTLs were also found to have environmental interaction effects.
017《Functional & Integrative Genomics》
Differential gene expression of rice two-component signaling elements during reproductive development and regulation by abiotic stress
Mukesh Jain, Akhilesh K. Tyagi, Jitendra P. Khurana
Interdisciplinary Centre for Plant Genomics and Department of Plant Molecular Biology, University of Delhi South Campus, Benito Juarez Road,
New Delhi, 110021, India
Abstract:The two-component signaling elements have been implicated in diverse cellular processes in plants. Earlier, we reported the identification,
characterization and expression analysis of type-A response regulators in rice. In this study, we have comprehensively analyzed the expression
profile of all the two-component signaling elements identified in rice at various stages of vegetative and reproductive development by employing
microarray analysis. Most of the components are expressed in all the developmental stages analyzed. A few of these were found to be specifically
expressed during certain stages of seed development, suggesting their role in embryo and endosperm development. In addition, some of these
components express differentially under various abiotic stress conditions, indicating their involvement at various levels of hierarchy in abiotic
stress signaling.
018《Euphytica》
Identification of QTLs controlling rice drought tolerance at seedling stage in hydroponic culture
Yoichiro Kato, Sawa Hirotsu, Keisuke Nemoto, Junko Yamagishi
Graduate School of Agricultural and Life Sciences, The University of Tokyo, Nishitokyo, Tokyo 188-0002, Japan
Abstract:'Drought avoidance' and 'drought tolerance' are two mechanisms by which plants adapt under water stress. These mechanisms are difficult
to evaluate separately in field experiments. Using hydroponic culture, we studied the genetic control of drought tolerance in rice (Oryza sativa L.)
without the effect of drought avoidance. A backcross inbred population of 'Akihikari' (lowland cultivar) × 'IRAT109' (upland cultivar) with 106
lines was cultured with (stressed condition) and without (non-stressed condition) polyethylene glycol (PEG) at seedling stage. The relative growth
rate (RGR), specific water use (SWU), and water use efficiency (WUE) showed significant genotype × environment interactions with or without
PEG, indicating that each line responded differently to water stress. A quantitative trait locus (QTL) analysis revealed that these interactions were
QTL specific. A total of three QTLs on chromosomes 2, 4, and 7 were detected for RGR. The QTL on chromosome 7 had a constant effect across
environments, while the QTL on chromosome 4 had an effect only under non-stressed condition and that on chromosome 2 only under stressed
condition. The stress-specific QTL on chromosome 2 was not co-located with any QTLs for root system depth previously reported from the same
mapping population. However, this QTL was co-located with a stress-specific QTL for SWU, suggesting that the control of transpiration was
relevant to dry matter production under drought. We concluded that PEG-treated hydroponic culture is very effective for use in genetic analyses of
drought tolerance at seedling stage.
019《Molecular Breeding》
The QTL controlling amino acid content in grains of rice ( Oryza sativa ) are co-localized with the regions involved in the amino acid metabolism
pathway
Lingqiang Wang, Ming Zhong, Xianghua Li, Dejun Yuan, Yunbi Xu, Huifang Liu, Yuqing He, Lijun Luo, Qifa Zhang
National Key Laboratory of Crop Genetic Improvement, National Center of Plant Gene Research and National Center of Crop Molecular Breeding,
Huazhong Agricultural University, Wuhan, 430070, China
Abstract:The improvement of grain quality, such as protein content (PC) and amino acid composition, has been a major concern of rice breeders.
We constructed a population of 190 recombinant inbred lines (RILs) from a cross between Zhenshan 97 and Nanyangzhan to map the quantitative
trait locus or loci (QTL) for amino acid content (AAC) as characterized by each of the AACs, total essential AAC, and all AAC. Using the data
collected from milled rice in 2002 and 2004, we identified 18 chromosomal regions for 19 components of AAC. For 13 of all the loci, the
Zhenshan 97 allele increased the trait values. Most QTL were co-localized, forming ten QTL clusters in 2002 and six in 2004. The QTL clusters
varied in both effects and locations, and the mean values of variation explained by individual QTL in the clusters ranged from 4.3% to 28.82%. A
relatively strong QTL cluster, consisting of up to 19 individual QTL, was found at the bottom of chromosome 1. The major QTL clusters identified
for two different years were coincident. A wide coincidence was found between the QTL we detected and the loci involved in amino acid
metabolism pathways, including N assimilation and transfer, and amino acid or protein biosynthesis. The results will be useful for candidate gene
identification and marker-assisted favorable allele transfer in rice breeding programs.
020《Plant Cell Reports》
Enhanced tolerance to drought stress in transgenic rice plants overexpressing a small heat-shock protein, sHSP17.7
Yutaka Sato, Sakiko Yokoya
National Agricultural Research Center for Hokkaido Region, Hitsujigaoka 1, Toyohira-ku, Sapporo 062-8555, Japan
Abstract:Exposure of rice (Oryza sativa L.) seedlings to a high temperature (42℃) for 24 h resulted in a significant increase in tolerance to drought
stress. To try to determine the mechanisms of acquisition of tolerance to drought stress by heat shock, the rice small heat-shock protein gene,
sHSP17.7, the product of which was shown to act as molecular chaperones in vitro and in vivo in our previous study, was overexpressed in the rice
cultivar “Hoshinoyume”. Western and Northern blot analyses showed higher expression levels of sHSP17.7 protein in three transgenic lines than in
one transgenic line. Drought tolerance was assessed in these transgenic lines and wild-type plants by withholding water for 6 days for evaluation of
the ability of plants to continue growth after water-stress treatments. Although no significant difference was found in water potential of seedlings
between transgenic lines and wild-type plants at the end of drought treatments, only transgenic seedlings with higher expression levels of
sHSP17.7 protein could regrow after rewatering. Similar results were observed in survival rates after treatments with 30% polyethylene glycol
(PEG) 3640 for 3 days. These results suggest that overproduction of sHSP17.7 could increase drought tolerance in transgenic rice seedlings.
021《Genetic Resources and Crop Evolution》
Reliable multiplication of seed for NERICA varieties of rice, Oryza sativa L.
Ryoichi Ikeda, Yoshimi Sokei, Inoussa Akintayo
African Rice Initiative: ARI/Africa Rice Center (WARDA), Cotonou, 01 BP 2031, Benin
Abstract:A comparison of breeder seed from different seed sources produced guidelines for multiplication of NERICA (New Rice for Africa)
varieties. Lower percentage detection of offtypes was noted with seed from a breeder than with seed from the Genetic Resources Unit (GRU) of
the Africa Rice Center. The GRU grows breeder seed in the field as bulk populations of varieties and this seed is multiplied for the foundation seed.
Breeders, however, always cultivate the variety in each family with several lines. In the comparison, each NERICA variety was cultivated as a
single plant per hill for easy offtype detection in the field. Any offtypes or doubtful plants were recorded and immediately discarded. The breeder
seed was not always genetically uniform because it was not yet fixed genetically and included mechanical seed mixture as well as natural mutants.
Offtypes were classified into categories such as chlorophyll mutants, aberrations of plant height, plant types and panicle types, semi- or high
sterility, awnness, apiculous color, leaf sheath color and excessively early plants. The causes of these offtypes were mainly (1) mechanical seed
mixture, (2) segregation in genetically-unfixed material, (3) out-crossing with another variety, or (4) natural mutation. In 2006, we multiplied
foundation seed using used home-saved seed from the 2005 crop harvested after removal of offtypes. The detection percentage of offtypes
decreased compared with the previous year, but the number of semi- or high sterility plants increased significantly. This review confirms that
breeder seed must be maintained with line culture within a family.
022《Plant Molecular Biology》
Overexpression of rice WRKY89 enhances ultraviolet B tolerance and disease resistance in rice plants
Haihua Wang, Junjie Hao, Xujun Chen, Zhongna Hao, Xia Wang, Yonggen Lou, Youliang Peng, Zejian Guo
Department of Plant Pathology, State Key Laboratory of Agrobiotechnology, China Agricultural University, Yuanmenyuan West Rd. 2, Beijing,
100094, China
Abstract:WRKY proteins are a large family of transcriptional regulators involved in a variety of biological processes in plants. Here we report
functional characterization of a rice WRKY gene, OsWRKY89. RNA gel blot analysis indicated that OsWRKY89 was strongly induced by
treatments of methyl jasmonate and UV-B radiation. The transient expression analysis of the OsWRKY89-eGFP reporter in onion epidermal cells
revealed that OsWRKY89 was targeted to nuclei. Transcriptional activity assays of OsWRKY89 and its mutants fused with a GAL4 DNA binding
domain indicated that the 67 C-terminal amino acids were required for the transcriptional activation and that the leucine zipper region at the
N-terminus enhanced its transcriptional activity. Overexpression of OsWRKY89 led to growth retardation at the early stage and reduction of
internode length. Scanning electron microscopy revealed an increase in wax deposition on leaf surfaces of the OsWRKY89 overexpression lines
and a decrease in wax loading in the RNAi-mediated OsWRKY89 suppression lines. Moreover, extractable and cell-wall-bound phenolic
compounds were decreased in the overexpressor lines, but its SA levels were increased. Lignin staining showed an increase in lignification in
culms of the overexpressor lines. Interestingly, overexpression of the OsWRKY89 gene enhanced resistance to the rice blast fungus and
white-backed planthopper as well as tolerance to UV-B irradiation. These results suggest that OsWRKY89 plays an important role in response to
biotic and abiotic stresses.
023《Planta》
New target for rice lodging resistance and its effect in a typhoon
Ken Ishimaru, Eiji Togawa, Taiichro Ookawa, Takayuki Kashiwagi, Yuka Madoka, Naoki Hirotsu
National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba Ibaraki, 305-8602, Japan
Abstract:We demonstrated the new target for lodging resistance in rice (Oryza sativa L.) by the analysis of physiological function of a locus for
lodging resistance in a typhoon (lrt5) with the near isogenic line under rice “Koshihikari” genetic background (tentatively named S1). The higher
lodging resistance of S1 was observed during a typhoon in September 2004 (28 days after heading), when most other plants in “Koshihikari”
became lodged. Visual observations showed that bending of the upper stems triggered lodging during the typhoon; the upper stem of “Koshihikari”
buckled completely, whereas that of S1 remained straight. In addition to the strong rain and winds during the typhoon, the weight of the buckled
upper plant parts increased the pressure on adjacent plants and caused a domino effect in “Koshihikari”. Young's modulus, an indicator of the
rigidity of the culm, was significantly higher in S1 than in “Koshihikari”. In the upper culm, the starch content in S1 was 4.8 times the value in
“Koshihikari”, and senescence was delayed in the upper leaves of S1. These results suggest that the rigidity of the upper culm by the higher starch
content (as a result of delayed senescence in the upper leaves) may be responsible for the higher lodging resistance during a typhoon in rice.
024《Molecular Genetics and Genomics》
MNU-induced mutant pools and high performance TILLING enable finding of any gene mutation in rice
Mamdou B. Barry, Jean Louis Pham, Sedou Béavogui, Alain Ghesquière, Nour Ahmadi
Institut de Recherche Agronomique de Guinée, PB 1523, Conakry, Guinea
Abstract:Greater insight into the dynamics of genetic resources of crop plants is needed in order to pinpoint detrimental evolutionary patterns and
draw up conservation priorities. Temporal evolution of rice genetic diversity was monitored in Maritime Guinea where subsistence-oriented
agriculture prevails. Diachronic comparison was performed between samples collected in six villages during the 1979/1982-period and in 2003,
based on the names and number of varieties inventoried and the polymorphism of microsatellite markers. The number of varieties appeared not to
be comparable between the two dates, due to differences in the collection methods. The varietal composition had evolved very substantially
between the two collection dates. Many long-duration varieties present in 1979/1982 had been abandoned and several improved varieties had been
introduced. The mean number of alleles per locus and per accession was significantly higher in accessions collected in 2003. Pairwise comparisons
of the mean number of alleles per locus in 1979/1982-2003 homonymous accession pairs indicated higher intra-accession diversity for the 2003
collections. Genetic differentiation, measured with the F ST values, was very high and significant for more than 80% of these pairs of accessions.
The overall genetic differentiation between accessions from the two collections dates was also significant. Significant changes were also observed
for allelic composition. However, alleles specific of each collection date had much lower frequency, compared to alleles common to the two
collection dates. These results suggest that rice genetic diversity in Maritime Guinea has been maintained or even enhanced. Old collections of
crop genetic resources are often not exhaustive enough to undertake perfect diachronic comparison. New methods to utilize this historical data for
diversity monitoring are needed.
025《The Plant Journal》
DWARF10, an RMS1/MAX4/DAD1 ortholog, controls lateral bud outgrowth in rice
Tomotsugu Arite, Hirotaka Iwata, Kenji Ohshima, Masahiko Maekawa, Masatoshi Nakajima, Mikiko Kojima, Hitoshi Sakakibara and Junko
Kyozuka
Graduate School of Agriculture and Life Sciences, University of Tokyo, Yayoi, Bunkyo, Tokyo 113-0032, Japan
Abstract:Plant architecture is mostly determined by shoot branching patterns. Apical dominance is a well-known control mechanism in the
development of branching patterns, but little is known regarding its role in monocots such as rice. Here, we show that the concept of apical
dominance can be applied to tiller bud outgrowth of rice. In dwarf10 (d10), an enhanced branching mutant of rice, apical dominance can be
observed, but the inhibitory effects of the apical meristem was reduced. D10 is a rice ortholog of MAX4/RMS1/DAD1 that encodes a carotenoid
cleavage dioxygenase 8 and is supposed to be involved in the synthesis of an unidentified inhibitor of shoot branching. D10 expression
predominantly occurs in vascular cells in most organs. Real-time polymerase chain reaction analysis revealed that accumulation of D10 mRNA is
induced by exogenous auxin. Moreover, D10 expression is upregulated in six branching mutants, d3, d10, d14, d17, d27 and high tillering dwarf
(htd1). No such effects were found for D3 or HTD1, the MAX2 and MAX3 orthologs, respectively, of rice. These findings imply that D10
transcription might be a critical step in the regulation of the branching inhibitor pathway. In addition, we present observations that suggest that
FINE CULM1 (FC1), a rice ortholog of teosinte branched 1 (tb1), possibly works independently of the branching inhibitor pathway.
026、
《The Plant Journal》
The rice nuclear gene, VIRESCENT 2, is essential for chloroplast development and encodes a novel type of guanylate kinase targeted to plastids
and mitochondria
Hiroki Sugimoto, Kensuke Kusumi, Ko Noguchi, Masahiro Yano, Atsushi Yoshimura and Koh Iba
Department of Biological Sciences, Faculty of Sciences, Kyushu University, Fukuoka 812-8581, Japan
Abstract:Guanylate kinase (GK) is a critical enzyme in guanine nucleotide metabolism pathways, catalyzing the phosphorylation of (d)GMP to
(d)GDP. Here we show that a novel gene, VIRESCENT 2 (V2), encodes a new type of GK (designated pt/mtGK) that is localized in plastids and
mitochondria. We initially identified the V2 gene by positional cloning of the rice v2 mutant. The v2 mutant is temperature-sensitive and develops
chlorotic leaves at restrictive temperatures. The v2 mutation causes inhibition of chloroplast differentiation; in particular, it disrupts the chloroplast
translation machinery during early leaf development [Sugimoto et al. (2004)Plant Cell Physiol. 45, 985]. In the bacterial and animal species
studied to date, GK is localized in the cytoplasm and participates in maintenance of the guanine nucleotide pools required for many fundamental
cellular processes. Phenotypic analysis of rice seedlings with RNAi knockdown of cytosolic GK (designated cGK) showed that cGK is
indispensable for the growth and development of plants, but not for chloroplast development. Thus, rice has two types of GK, as does Arabidopsis,
suggesting that higher plants have two types of GK. Our results suggest that, of the two types of GK, only pt/mtGK is essential for chloroplast
differentiation.
027《Euphytica》
Identification of near-isogenic lines: an innovative approach, validated for root and shoot morphological characters in a mapping population of rice
( Oryza sativa L.)
H. R. Prabuddha, K. Manjunatha, R. Venuprasad, M. S. Vinod, J. H. Jureifa, H. E. Shashidhar
Marker-Assisted Selection Laboratory, Department of Genetics and Plant Breeding, College of Agriculture, University of Agricultural Sciences,
GKVK, Bangalore, 560065, India
Abstract:Near-isogenic lines (NILs) constitute valuable tools in genetic investigations and plant breeding programs. Conventional methods for
developing these are time consuming and tedious. An innovative method for identifying NILs is proposed and validated. The method involves
computation of simple correlation coefficients of all possible pairs of genotypes within a mapping population using molecular marker data, and
phenotypic characterization of those pairs with very high positive correlation. The pairs having both genomic and phenotypic similarity except for
a single trait are considered as NILs. This strategy was tested with a doubled haploid mapping population involving CT9993 and IR62266. This
population was saturated with 315 markers and comprised 154 lines. The pairs showing very high correlation coefficients (0.70-0.97) and differing
for less than 10% of the markers were considered as Genotypically Closely Related Pairs (GCRPs). Graphical genotyping was employed to
visualize the genome of the closely related lines. A total of 39 such pairs were subjected to rigorous evaluation for root and shoot morphological
traits in two contrasting moisture regimes. Four GCRPs under well-watered condition and ten GCRPs under low moisture stress condition are
statistically significant for a single phenotypic trait and are considered as NILs for their respective traits and would be the valuable materials for
genetic studies. Mapped QTLs and candidate genes were employed to explain the probable cause of phenotypic difference in NILs.
028《Plant Cell Reports》
Genetic transformation of NERICA, interspecific hybrid rice between Oryza glaberrima and O. sativa , mediated by Agrobacterium tumefaciens
Takuma Ishizaki, Takashi Kumashiro
Biological Resources Division, Japan International Research Center for Agricultural Sciences (JIRCAS), 1-1 Ohwashi, Tsukuba Ibaraki, 305-8686,
Japan
Abstract:We developed an efficient gene transfer method mediated by Agrobacterium tumefaciens for introgression of new rice for Africa
(NERICA) cultivars, which are derivatives of interspecific hybrids between Oryza glaberrima Steud. and O. sativa L. Freshly isolated immature
embryos were inoculated with A. tumefaciens LBA4404 that harbored binary vector pBIG-ubi::GUS or pIG121Hm, which each carried a
hygromycin-resistance gene and a GUS gene. Growth medium supplemented with 500 mg/l cefotaxime and 20 mg/l hygromycin was suitable for
elimination of bacteria and selection of transformed cells. Shoots regenerated from the selected cells on MS medium containing 20 g/l sucrose, 30
g/l sorbitol, 2 g/l casamino acids, 0.25 mg/l naphthaleneacetic acid, 2.5 mg/l kinetin, 250 mg/l cefotaxime, and 20 mg/l hygromycin. The shoots
developed roots on hormone-free MS medium containing 30 mg/l hygromycin. Integration and expression of the transgenes were confirmed by
PCR, Southern blot analysis, and histochemical GUS assay. Stable integration, expression, inheritance, and segregation of the transgenes were
demonstrated by molecular and genetic analyses in the T0 and T1 generations. Most plants were normal in morphology and fertile. The
transformation protocol produced stable transformants from 16 NERICA cultivars. We also obtained transformed plants by inoculation of calluses
derived from mature seeds, but the frequency of transformation was lower and sterility was more frequent.
029《Planta》
Identification of the OsOPR7 gene encoding 12-oxophytodienoate reductase involved in the biosynthesis of jasmonic acid in rice
Tomoyuki Tani, Hiroyuki Sobajima, Kazunori Okada, Tetsuya Chujo, Shin-ichi Arimura, Nobuhiro Tsutsumi, Mikio Nishimura, Hideharu Seto,
Hideaki Nojiri, Hisakazu Yamane
Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
Abstract:Enzyme 12-oxophytodienoate (OPDA) reductase (EC1.3.1.42), which is involved in the biosynthesis of jasmonic acid (JA), catalyses the
reduction of 10, 11-double bonds of OPDA to yield 3-oxo-2-(2'-pentenyl)-cyclopentane-1-octanoic acid (OPC-8:0). The rice OsOPR1 gene
encodes OPDA reductase (OPR) converting (−)-cis-OPDA preferentially, rather than (+)-cis-OPDA, a natural precursor of JA. Here, we provide
evidence that an OPR family gene in rice chromosome 8, designated OsOPR7, encodes the enzyme involved in the JA biosynthesis. Recombinant
OsOPR7-His protein efficiently catalysed the reduction of both enantiomers of cis-OPDA, similar to the OPR3 protein in Arabidopsis thaliana (L.)
Heynh. The expression of OsOPR7 mRNA was induced and reached maximum levels within 0.5 h of mechanical wounding and drought stress,
and the endogenous JA level started to increase in accordance with the increase in OsOPR7 expression. The GFP-OsOPR7 fusion protein was
detected exclusively in peroxisomes in onion epidermal cells. Furthermore, complementation analysis using an Arabidopsis opr3 mutant indicated
that the OsOPR7 gene, but not OsOPR1, was able to complement the phenotypes of male sterility in the mutant caused by JA deficiency, and that
JA production in the opr3 mutant was also restored by the expression of the OsOPR7 gene. We conclude that the OsOPR7 gene encodes the
enzyme catalysing the reduction of natural (+)-cis-OPDA for the JA biosynthesis in rice.
030《Planta》
Cation dependent O -methyltransferases from rice
Yoon Jung Lee, Bong Gyu Kim, Youhoon Chong, Yoongho Lim, Joong-Hoon Ahn
Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, Seoul, 143-701, South Korea
Abstract:Two lower molecular mass OMT genes (ROMT-15 and -17) were cloned from rice and expressed in Escherichia coli as glutathione
S-transferase fusion proteins. ROMT-15 and -17 metabolized caffeoyl-CoA, flavones and flavonols containing two vicinal hydroxyl groups,
although they exhibited different substrate specificities. The position of methylation in both luteolin and quercetin was determined to be the 3'
hydroxyl group and myricetin and tricetin were methylated not only at 3' but also at 5' hydroxyl groups. ROMT-15 and -17 are cation-dependent
and mutation of the predicted metal binding sites resulted in the loss of the enzyme activity, indicating that the metal ion has a critical role in the
enzymatic methylation.
031《Biochemical Genetics》
Abundant Within-varietal Genetic Diversity in Rice Germplasm from Yunnan Province of China Revealed by SSR Fingerprints
Min Tu, Bao-Rong Lu, Youyong Zhu, Yunyue Wang
Ministry of Education Key Laboratory of Agricultural Biodiversity for Plant Disease Management, Key Laboratory of Plant Pathology, Yunnan
Agricultural University, Kunming, 650201, P.R. China
Abstract:In order to estimate genetic diversity of rice (Oryza sativa L.) germplasm in Yunnan Province of China, 60 varieties from different regions
were analyzed by microsatellite (SSR) fingerprints. Nine selected SSR primer pairs amplified a total of 55 alleles from these varieties, and high
genetic diversity (0.706) was found, although it was not evenly distributed across the regions. Marked genetic variation was detected within the
traditional varieties. A UPGMA dendrogram based on SSR polymorphism indicated a great variation among the rice varieties, with coefficients
ranging between 0.229 and 1.000. The formation of the rice diversity pattern in Yunnan is associated with natural conditions and especially with
diverse cultural demands and farming styles. Strategic conservation of rice germplasm in Yunnan is important, and this could be implemented by
collecting varieties across geographic regions with sufficient individuals within the same varieties. Effective rice conservation should also consider
cultural aspects during collection.
032《Molecular Breeding》
Inheritance and linkage analysis of fertility restoration genes for WA, Dissi, and Gambiaca cytoplasmic male sterility systems in rice
Majid Sattari, Arumugam Kathiresan, Glenn B. Gregorio, Sant S. Virmani
Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute (IRRI), DAPO Box 7777, Metro Manila, Philippines
Abstract:The genetic relationship among three cytoplasmic male sterility (CMS) systems—wild abortive (WA), Dissi, and Gambiaca—was studied
using the parents of well performing rice hybrids. The results showed that maintainers of one CMS system can also maintain sterility in other
cytoplasmic backgrounds. F1 plants derived from crosses involving A and R lines of the respective cytoplasm and combinations of their crosses
with other CMS systems showed similar pollen and spikelet fertility values, indicating that similar biological processes govern fertility restoration
in these three CMS systems. The results of inheritance studies showed that pollen fertility restoration in all three CMS systems is governed by two
independent and dominant genes with classical duplicate gene action. Three F 2 populations, generated from crosses of IR58025A/IR34686R (WA),
IR75596A/IR68077-82-22-2-3R (Dissi), and IR75601A/IR68444R (Gambiaca), were used to map the Rf genes. For the WA-CMS system, Rf3
was located at a distance of 2.8 cM from RM490 on chromosome 1, and Rf4 was located at 1.6 cM from RM1108 on chromosome 10. For the
Dissi-CMS system, Rf3 was located on chromosome 1 at 1.9 cM from RM7466, and Rf4 on chromosome 10 was located at 2.3 cM from RM6100.
The effect of Rf3 on pollen fertility appeared to be stronger than that of Rf4. In the Gambiaca-CMS system, only one major locus was mapped on
chromosome 1 at 2.1 cM from RM576. These studies have led to the development of PCR-based markers for marker-assisted selection (MAS) for
selecting putative restorer lines, new approaches to alloplasmic line breeding, and the transfer of Rf genes into adapted cultivars through a
backcrossing program in an active hybrid rice breeding program.
033《Plant Molecular Biology》
A genome-wide gain-of-function analysis of rice genes using the FOX-hunting system
Hidemitsu Nakamura, Makoto Hakata, Kou Amano, Akio Miyao, Naoko Toki, Mariko Kajikawa, Jinhuan Pang, Naokuni Higashi, Shigeko Ando,
Seiichi Toki, Miki Fujita, Akiko Enju, Motoaki Seki, Miki Nakazawa, Takanari Ichikawa, Kazuo Shinozaki, Minami Matsui, Yoshiaki Nagamura,
Hirohiko Hirochika, Hiroaki Ichikawa
/Division of Genome and Biodiversity Research, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba Ibaraki, 305-8602,
Japan
Abstract:The latest report has estimated the number of rice genes to be ∼32 000. To elucidate the functions of a large population of rice genes and
to search efficiently for agriculturally useful genes, we have been taking advantage of the Full-length cDNA Over-eXpresser (FOX) gene-hunting
system. This system is very useful for analyzing various gain-of-function phenotypes from large populations of transgenic plants overexpressing
cDNAs of interest and others with unknown or important functions. We collected the plasmid DNAs of 13 980 independent full-length cDNA
(FL-cDNA) clones to produce a FOX library by placing individual cDNAs under the control of the maize Ubiquitin-1 promoter. The FOX library
was transformed into rice by Agrobacterium-mediated high-speed transformation. So far, we have generated approximately 12 000 FOX-rice lines.
Genomic PCR analysis indicated that the average number of FL-cDNAs introduced into individual lines was 1.04. Sequencing analysis of the PCR
fragments carrying FL-cDNAs from 8615 FOX-rice lines identified FL-cDNAs in 8225 lines, and a database search classified the cDNAs into
5462 independent ones. Approximately 16.6% of FOX-rice lines examined showed altered growth or morphological characteristics. Three
super-dwarf mutants overexpressed a novel gibberellin 2-oxidase gene, confirming the importance of this system. We also show here the other
morphological alterations caused by individual FL-cDNA expression. These dominant phenotypes should be valuable indicators for gene discovery
and functional analysis.
034《TAG Theoretical and Applied Genetics》
A novel gene, Pi40(t) , linked to the DNA markers derived from NBS-LRR motifs confers broad spectrum of blast resistance in rice
J. U. Jeung, B. R. Kim, Y. C. Cho, S. S. Han, H. P. Moon, Y. T. Lee, K. K. Jena
Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute, Los Baños, Laguna, Philippines
Abstract:Rice blast disease caused by Magnaporthe grisea is a continuous threat to stable rice production worldwide. In a modernized agricultural
system, the development of varieties with broad-spectrum and durable resistance to blast disease is essential for increased rice production and
sustainability. In this study, a new gene is identified in the introgression line IR65482-4-136-2-2 that has inherited the resistance gene from an EE
genome wild Oryza species, O. australiensis (Acc. 100882). Genetic and molecular analysis localized a major resistance gene, Pi40(t), on the short
arm of chromosome 6, where four blast resistance genes (Piz, Piz-5, Piz-t, and Pi9) were also identified, flanked by the markers S2539 and
RM3330. Through e-Landing, 14 BAC/PAC clones within the 1.81-Mb equivalent virtual contig were identified on Rice Pseudomolecule3. Highly
stringent primer sets designed for 6 NBS-LRR motifs located within PAC clone P0649C11 facilitated high-resolution mapping of the new
resistance gene, Pi40(t). Following association analysis and detailed haplotyping approaches, a DNA marker, 9871.T7E2b, was identified to be
linked to the Pi40(t) gene at the 70 Kb chromosomal region, and differentiated the Pi40(t) gene from the LTH monogenic differential lines
possessing genes Piz, Piz-5, Piz-t, and Pi-9. Pi40(t) was validated using the most virulent isolates of Korea as well as the Philippines, suggesting a
broad spectrum for the resistance gene. Marker-assisted selection (MAS) and pathotyping of BC progenies having two japonica cultivar genetic
backgrounds further supported the potential of the resistance gene in rice breeding. Our study based on new gene identification strategies provides
insight into novel genetic resources for blast resistance as well as future studies on cloning and functional analysis of a blast resistance gene useful
for rice improvement.
035《In Vitro Cellular & Developmental Biology - Plant》
Development and characterization of a chimaeric tissue-specific promoter in wheat and rice endosperm
Maria Oszvald, Mark Gardonyi, Cecília Tamas, Imre Takacs, Barnabas Jenes, Laszlo Tamas
Department of Biochemistry and Food Technology, BUTE University, Budapest, Hungary
Abstract:The recently achieved significant improvement of cereal transformation protocols provides facilities to alter the protein composition of
the endosperm, for example, to increase or decrease the quantity of one of its protein components or to express foreign molecules. To achieve this
goal, strong endosperm-specific promoters have to be available. The aim of our work was to develop a more efficient tissue-specific promoter
which is currently used. A chimaeric promoter was assembled using the 5' UTR (1,900 bp) of the gene coding for the 1Bx17 HMW glutenin
subunit protein, responsible for tissue-specific expression and the first intron of the rice actin gene (act1). The sequence around of the translation
initial codon was optimized. The effect of the intron and promoter regulatory sequences, using different lengths of 1Bx17 HMW-GS promoter,
were studied on the expression of uidA gene. The function of promoter elements, promoter length, and the first intron of the rice actin gene were
tested by a transient expression assay in immature wheat endosperm and in stable transgenic rice plants. Results showed that insertion of the rice
act1 first intron increased GUS expression by four times in transient assay. The shortest 1Bx17 HMW-GS promoter fragment (173 bp) linked to
the intron and GUS reporter gene provided almost the same expression level than the intronless long 1Bx17 HMW-GS promoter. Analysis of the
stable transformant plants revealed that 173 nucleotides were sufficient for endosperm-specific expression of the uidA gene, despite 13 nucleotides
missing from the HMW enhancer sequence, a relevant regulatory element in the promoter region.
036《Theoretical and Applied Genetics》
Evaluation of genes from eIF4E and eIF4G multigenic families as potential candidates for partial resistance QTLs to Rice yellow mottle virus in
rice
Arnaud Boisnard, Laurence Albar, Deless Thiéméle, Myriam Rondeau, Alain Ghesquière
UMR 5096, Laboratoire Génome et Développement des Plantes, IRD/CNRS/Université de Perpignan, BP 64501, 34394 Montpellier CEDEX 5,
France
Abstract:QTLs for partial resistance to Rice yellow mottle virus (RYMV) in rice were mapped in two populations of doubled-haploid lines (DHLs)
and recombinant inbred lines (RILs) derived from the same cross but evaluated for different resistance criteria (virus content and symptom
severity). An integrative map was used to compare the two genetic maps and a global analysis of both populations was performed. Most of the
QTLs previously identified in DHL population were confirmed with increased significance and precision. As many recent studies evidenced the
role of eukaryotic translation initiation factors (eIF) of 4E and 4G families in plant susceptibility to RNA viruses, we checked if these genes
co-locate with QTLs of resistance to RYMV. Their systematic in silico identification was carried out on the rice genome and their physical
locations were compared to QTL positions on the integrative map. In order to confirm or not the co-locations observed, the analysis was completed
by evaluation of near-isogenic lines, QTL fine mapping and sequencing of candidate genes. Three members from eIF4G family could be retained
as reliable candidates whereas eIF4E genes, commonly found to govern resistances in other plant/virus interactions, were discarded. Together with
the recent identification of an eIF(iso)4G as a major resistance gene, data suggests an important role of genes from eIF4G family in rice resistance
to RYMV but does not exclude the contribution of factors different from the translation initiation complex.
037《European Journal of Plant Pathology
Chitinase levels in rice cultivars correlate with resistance to the sheath blight pathogen Rhizoctonia solani
C. L. Shrestha, I. Oña, S. Muthukrishnan, T. W. Mew
Entomology and Plant Pathology Division, International Rice Research Institute (IRRI), Los Baños, Laguna, Philippines
Abstract:Various rice cultivars were selected and screened for their reaction to sheath blight in the greenhouse. Cluster analysis of percent relative
lesion height (% RLH) generated four groups of cultivars with a coefficient of similarity of 3.27. Chitinase activities were detected 24 h after
inoculation of moderately resistant cvs Betichikon, Dudruchi, Khatochalani, Padi Pulut Malat, Kakua, IR72, Khakibinni. But in the susceptible cv.
IR58, chitinase activity was detected only 36 h after inoculation. Western blot analysis showed that class 1 and class 2 chitinases were induced
following Rhizoctonia solani infection of these cultivars. The % RLH and the number of infection cushions were negatively correlated with the
level of chitinase activity. Moderately resistant rice cultivars had higher levels of chitinase activity and lower disease severity and numbers of
infection cushions formed compared to IR58.
038《Plant Molecular Biology》
Ds insertion mutagenesis as an efficient tool to produce diverse variations for rice breeding
Shu-Ye Jiang, Doris Bachmann, Honggui La, Zhigang Ma, Prasanna Nori Venkatesh, Rengasamy Ramamoorthy, Srinivasan Ramachandran
Rice Functional Genomics Group, Temasek Life Sciences Laboratory, 1 Research Link, The National University of Singapore, Singapore, 117604,
Singapore
Abstract:The availability of diversified germplasm resources is the most important for developing improved rice varieties with higher seed yield or
tolerance to various biotic or abiotic stresses. Here we report an efficient tool to create increased variations in rice by maize Ac/Ds transposon (a
gene trap system) insertion mutagenesis. We have generated around 20,000 Ds insertion rice lines of which majority are homozygous for Ds
element. We subjected these lines to phenotypic and abiotic stress screens and evaluated these lines with respect to their seed yields and other
agronomic traits as well as their tolerance to drought, salinity and cold. Based on this evaluation, we observed that random Ds insertions into rice
genome have led to diverse variations including a range of morphological and conditional phenotypes. Such differences in phenotype among these
lines were accompanied by differential gene expression revealed by GUS histochemical staining of gene trapped lines. Among the various
phenotypes identified, some Ds lines showed significantly higher grain yield compared to wild-type plants under normal growth conditions
indicating that rice could be improved in grain yield by disrupting certain endogenous genes. In addition, several 1,000s of Ds lines were subjected
to abiotic stresses to identify conditional mutants. Subsequent to these screens, over 800 lines responsive to drought, salinity or cold stress were
obtained, suggesting that rice has the genetic potential to survive under abiotic stresses when appropriate endogenous genes were suppressed. The
mutant lines that have higher seed yielding potential or display higher tolerance to abiotic stresses may be used for rice breeding by conventional
backcrossing combining with molecular marker-assisted selection. In addition, by exploiting the behavior of Ds to leave footprints upon
remobilization, we have shown an alternative strategy to develop new rice varieties without foreign DNA sequences in their genome.
039《Euphytica》
Marker assisted introgression of bacterial blight resistance in Samba Mahsuri, an elite indica rice variety
Raman M. Sundaram, Manne R. Vishnupriya, Sunil K. Biradar, Gouri S. Laha, Gajjala Ashok Reddy, N. Shobha Rani, Nukala P. Sarma, Ramesh
Venkata Sonti
Directorate of Rice Research, Rajendranagar, Hyderabad, 500030, India
Abstract:Samba Mahsuri (BPT5204) is a medium slender grain indica rice variety that is very popular with farmers and consumers across India
because of its high yield and excellent cooking quality. However, the variety is susceptible to several diseases and pests, including bacterial blight
(BB). We have used PCR based molecular markers in a backcross-breeding program to introgress three major BB resistance genes (Xa21, xa13
and xa5) into Samba Mahsuri from a donor line (SS1113) in which all the three genes are present in a homozygous condition. At each backcross
generation, markers closely linked to the three genes were used to select plants possessing these resistance genes (foreground selection) and
microsatellite markers polymorphic between donor and recurrent parent were used to select plants that have maximum contribution from the
recurrent parent genome (background selection). A selected BC4F1 plant was selfed to generate homozygous BC4F2 plants with different
combinations of BB resistance genes. The three-gene pyramid and two-gene pyramid lines exhibited high levels of resistance against the BB
pathogen. Under conditions of BB infection, the three-gene pyramid lines exhibited a significant yield advantage over Samba Mahsuri. Most
importantly, these lines retain the excellent grain and cooking qualities of Samba Mahsuri without compromising the yield as determined in
multi-location trials. This work demonstrates the successful application of marker-assisted selection for targeted introgression of multiple
resistance genes into a premium quality rice variety.
040《Molecular Breeding》
Genomic characterization of Oryza species-specific CACTA-like transposon element and its application for genomic fingerprinting of rice
varieties
Hee-Wan Kang, Kwon-Kyoo Kang
Graduate School of Biotechnology and Information Technology, Hankyong National University, Ansung, 456-749, Korea
Abstract:A repeated DNA fragment (pKRD) was isolated from the genomic library of weedy rice in Korea. The pKRD showed significant
homology to Em/Spm CACTA-like transposon in whole genome sequences of rice released in the Blast rice sequence database of NCBI and was
closely related to the TNP2 transposase group, including a TNP-like transposable element of rice. A Southern hybridization experiment
demonstrated that the pKRD sequence is unique to the Oryza genome. The 126 sequences homologous to pKRD were evenly distributed in all 12
different chromosomes in rice genomes with multiple copy numbers. Different copy numbers ranging from 1,500 to 4,500 corresponding to rice
species were detected by slot blot hybridization. In a DNA fingerprinting experiment, a pKRD probe was assessed to be the potential molecular
marker for studying evolution and divergence, biodiversity and phylogenic analysis of rice species.
041《Theoretical and Applied Genetics》
The number of genes having different alleles between rice cultivars estimated by SNP analysis
Kenta Shirasawa, Hiroaki Maeda, Lisa Monna, Sachie Kishitani, Takeshi Nishio
Graduate School of Agricultural Science, Tohoku University, Aoba-ku, Sendai 981-8555, Japan
Abstract:Identification of single nucleotide polymorphisms (SNPs) in a large number of genes will enable estimation of the number of genes
having different alleles in a population. In the present study, SNPs between 21 rice cultivars including 17 Japanese cultivars, one upland rice, and
three indica cultivars were analyzed by PCR-RF-SSCP. PCR-RF-SSCP analysis was found to be a more efficient method for detecting SNPs than
mismatch-cleavage analysis, though both PCR-RF-SSCP and mismatch-cleavage are useful for screening SNPs. The number of DNA fragments
showing polymorphism between Japanese cultivars was 134 in the 1,036 genes analyzed. In 137 genes, 638 DNA polymorphisms were identified.
Out of 52 genes having polymorphisms in the exons, one had a frame-shift mutation, three had polymorphism causing amino acid insertions or
deletions, and 16 genes had missense polymorphisms. The number of genes having frame-shift mutations and missense polymorphisms between
the 17 Japanese cultivars was estimated to be 41 and 677 on average, respectively, and those between japonica and indica to be 425 and 6,977,
respectively. Chromosomal regions of cultivars selected in rice breeding processes were identified by SNP analysis of genes.
042《Molecular Genetics and Genomics》
Characterization of two rice peroxidase promoters that respond to blast fungus-infection
Katsutomo Sasaki, Ohtsu Yuichi, Susumu Hiraga, Yoko Gotoh, Shigemi Seo, Ichiro Mitsuhara, Hiroyuki Ito, Hirokazu Matsui, Yuko Ohashi
Division of Plant Sciences, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8602, Japan
Abstract:Peroxidase (POX) genes consist of a large gene family possibly contributing to self-defense, however constitutive and stress-induced
expression patterns of individual gene were poorly understood in rice. We studied here the characteristic expression of two representative rice POX
genes, R2329 and R2184, which are blast fungus-inducible (Sasaki et al. in Plant Cell Physiol 45:1442-1452, 2004). Basal GUS activity in R2329
promoter::GUS rice plants was 100-fold higher than that in R2184 promoter::GUS plants, and these levels reflected the transcript levels monitored
by quantitative real-time RT-PCR. R2329 promoter was activated by blast fungus-infection and wounding, and R2184 promoter was activated by
the fungal-infection and methyl jasmonate (MeJA)-treatment. By histochemical GUS staining analysis, constitutive R2329 and R2184 expression
was commonly found in vascular bundle and exodermis in leaves and roots, while the precise expression profile was characteristic. In blast fungus
inoculated R2329 promoter::GUS leaves, GUS staining was induced just around fungus-induced local lesions. Analysis of the 5' deleted promoters
suggests the presence of many kinds of stress-responsive elements in the regions between −1798 and −748 of R2329 promoter and between −1975
and −548 of R2184 promoter. These results revealed the stress-responsive characteristics of R2329 and R2184 promoters, and indicated the
possible use for generation of useful transgenic plants.
043《Placenta》
Characterization of rice tryptophan decarboxylases and their direct involvement in serotonin biosynthesis in transgenic rice
Sei Kang, Kiyoon Kang, Kyungjin Lee, Kyoungwhan Back
Department of Molecular Biotechnology, Agricultural Plant Stress Research Center, Biotechnology Research Institute, Chonnam National
University, Gwangju, 500-757, South Korea
Abstract:l-Tryptophan decarboxylase (TDC) and l-tyrosine decarboxylase (TYDC) belong to a family of aromatic l-amino acid decarboxylases and
catalyze the conversion of tryptophan and tyrosine into tryptamine and tyramine, respectively. The rice genome has been shown to contain seven
TDC or TYDC-like genes. Three of these genes for which cDNA clones were available were characterized to assign their functions using
heterologous expression in Escherichia coli and rice (Oryza sativa cv. Dongjin). The purified products of two of the genes were expressed in E.
coli and exhibited TDC activity, whereas the remaining gene could not be expressed in E. coli. The recombinant TDC protein with the greatest
TDC activity showed a K m of 0.69 mM for tryptophan, and its activity was not inhibited by phenylalanine or tyrosine, indicating a high level of
substrate specificity toward tryptophan. The ectopic expression of the three cDNA clones in rice led to the abundant production of the products of
the encoded enzymes, tyramine and tryptamine. The overproduction of TYDC resulted in stunted growth and a lack of seed production due to
tyramine accumulation, which increased as the plant aged. In contrast, transgenic plants that produced TDC showed a normal phenotype and
contained 25-fold and 11-fold higher serotonin in the leaves and seeds, respectively, than the wild-type plants. The overproduction of either
tyramine or serotonin was not strongly related to the enhanced synthesis of tyramine or serotonin derivatives, such as feruloyltyramine and
feruloylserotonin, which are secondary metabolites that act as phytoalexins in plants.
044《Plant Cell Reports》
Increased senescence-associated gene expression and lipid peroxidation induced by iron deficiency in rice roots
Raul Antonio Sperotto, Tatiana Boff, Guilherme Leitão Duarte, Janette Palma Fett
Centro de Biotecnologia, Universidade Federal do Rio Grande do Sul, P.O. Box 15005, 91501-970 Porto Alegre, RS, Brazil
Abstract:Iron deficiency is among the most common nutritional disorders in plants. Low iron supply causes decreased root growth and even plant
death. However, there are no reports about the specific pathways that lead Fe-deficient roots to senescence and death. To investigate the molecular
mechanisms that regulate rice roots response to Fe-deficiency, rice seedlings were grown for 3, 6 and 9 days in the presence or absence of Fe.
Sequences of 28 induced genes in rice roots under Fe-deficiency were identified by representational difference analysis (RDA). About 40% of
these sequences have been previously reported as senescence-related. Differential expression of selected genes was confirmed by semi-quantitative
RT-PCR analysis. Classical senescence-related sequences, such as MYB and WRKY transcription factors, cysteine protease, ubiquitin-conjugating
enzyme, lipid transfer protein, fatty acid hydroxylase, β-glucosidase and cytochrome P450 oxydoreductase were identified. Fe-deficiency also
resulted in decreased dry weight, increased lipid peroxidation (detected by TBA and histochemical methods) as well as evident membrane damage
in Fe-deficient roots. Taken together, the results indicate that Fe-deficiency in roots is linked to typical senescence pathways, associated with lipid
peroxidation.
045《Plant Molecular Biology》
Downstream promoter sequence of an Indian isolate of Rice tungro bacilliform virus alters tissue-specific expression in host rice and acts
differentially in heterologous system
Saloni Mathur, Indranil Dasgupta
Department of Plant Molecular Biology, University of Delhi South Campus, Benito Juarez Road, New Delhi, 110021, India
Abstract:An Indian isolate of Rice tungro bacilliform virus from West Bengal (RTBV-WB) showed significant nucleotide differences in its putative
promoter region when compared with a previously characterized isolate from Philippines. The transcription start site of RTBV-WB was mapped
followed by assessing the activity and tissue-specificity of the full-length (FL) promoter (−231 to +645) and several of its upstream and
downstream deletions by studying the expression of β-Glucuronidase (GUS) reporter gene in transgenic rice (Oryza sativa L. subsp. indica) plants
at various stages of development. In addition to the expected vascular-specific expression pattern, studied by histochemical staining, GUS
enzymatic assay and northern and RT-PCR analysis, two novel patterns were revealed in some of the downstream deleted versions; a
non-expressing type, representing no expression at any stage in any tissue and constitutive type, representing constitutive expression at all stages in
most tissues. This indicated the presence of previously unreported positive and negative cis-regulatory elements in the downstream region. The
negative element and a putative enhancer region in the upstream region specifically bound to rice nuclear proteins in vitro. The FL and its deletion
derivatives were also active in heterologous systems like tobacco (Nicotiana tabacum) and wheat (Triticum durum). Expression patterns in tobacco
were different from those observed in rice suggesting the importance of upstream elements in those systems and host-specific regulation of the
promoter in diverse organisms. Thus, the RTBV-WB FL promoter and its derivatives contain an array of cis-elements, which control constitutive or
tissue- and development-specific gene expression in a combinatorial fashion.
046《Plant Cell Reports》
Knockout of a starch synthase gene OsSSIIIa / Flo5 causes white-core floury endosperm in rice ( Oryza sativa L.)
Nayeon Ryoo, Chul Yu, Cheon-Seok Park, Moo-Yeol Baik, In Myoung Park, Man-Ho Cho, Seong Hee Bhoo, Gynheung An, Tae-Ryong Hahn,
Jong-Seong Jeon
Plant Metabolism Research Center and Graduate School of Biotechnology, Kyung Hee University, Yongin , 446-701, Korea
Abstract:To elucidate the role of SSIIIa during starch synthesis in rice (Oryza sativa L.) endosperm, we characterized null mutants of this gene,
generated by T-DNA insertions. Scanning electron microscope (SEM) analysis revealed that the starch granules in these mutants are smaller and
rounder compared with the wild type controls, and that the mutant endosperm is characterized by a loosely packed central portion exhibiting a
floury-like phenotype. Hence, the OsSSIIIa (Oryza sativa SSIIIa) mutations are referred to as white-core floury endosperm 5-1 (flo5-1) and flo5-2.
Based upon their X-ray diffraction patterns, the crystallinity of the starch in the flo5 mutant endosperm is decreased compared with wild type.
Through determination of the chain-length distribution of the mutant endosperm starch, we found that flo5-1 and flo5-2 mutants have reduced the
content of long chains with degree of polymerization (DP) 30 or greater compared with the controls. This suggests that OsSSIIIa/Flo5 plays an
important role in generating relatively long chains in rice endosperm. In addition, DP 6 to 8 and DP 16 to 20 appeared to be reduced in endosperm
starch of flo5-1 and flo5-2, whereas DP 9 to 15 and DP 22 to 29 were increased in these mutants. By the use of differential scanning calorimetry
(DSC), the gelatinization temperatures of endosperm starch were found to be 1–5°C lower than those of the control. We propose a distinct role for
OsSSIIIa/Flo5 and the coordinated action of other SS isoforms during starch synthesis in the seed endosperm of rice.
047《Conservation Genetics》
Genetic structure of three Oryza AA genome species (O. rufipogon, O. nivara and O. sativa) as assessed by SSR analysis on the Vientiane Plain of
Laos
Yosuke Kuroda, Yo-Ichiro Sato, Chay Bounphanousay, Yasuyuki Kono, Koji Tanaka
Graduate School of Asian and African Area Studies, Kyoto University Sakyo-ku, Kyoto 603-8501, Japan
Abstract:Microsatellite (SSR) markers can reveal a high level of polymorphic loci, and are increasingly being used in population genetic structure
studies. On the Vientiane plain of Laos all components of the rice crop complex exist, wild annual (O. nivara), wild perennial (O. rufipogon) and
weedy relatives of rice as well as rice itself. To understand gene flow in the rice complex, the genetic structures of O. rufipogon (10 populations),
O. nivara (10 populations) and O. sativa (24 samples) from across the Vientiane Plain, Laos, were compared. Higher genetic differentiation was
detected among O. nivara populations (G ST = 0.77, R ST = 0.71) than O. rufipogon populations (G ST = 0.29, R ST = 0.28), whereas genetic
diversity for all populations of these two wild species showed similar values (H T = 0.77 and 0.64 in O. rufipogon and O. nivara, respectively).
Based on neighbor-joining tree constructed on the basis of genetic distance (D A), three genetic clusters were detected, corresponding to (1) O.
sativa samples, (2) O. nivara populations and (3) O. rufipogon populations. Pairwise tests confirmed the genetic differentiation of the three species.
Although none of the wild rice individuals used in this study had any cultivated-specific phenotypic traits, genetic admixture analysis detected
more than 10% O. sativa membership in three O. rufipogon and one O. nivara populations, indicating that O. sativa alleles may cryptically persist
in natural populations of O. rufipogon and O. nivara on the Vientiane Plain.
048《Euphytica》
Molecular diversity and genome-wide linkage disequilibrium patterns in a worldwide collection of Oryza sativa and its wild relatives
H. A. Agrama, G. C. Eizenga
Rice Research and Extension Center, University of Arkansas, Stuttgart, AR 72160, USA
Abstract:Genetic diversity analysis within a species is vital for understanding evolutionary processes at the population and genomic levels. We
report a detailed study of molecular diversity, polymorphism and linkage disequilibrium in three groups of rice (Oryza) germplasm accessions
based on 176 SSR markers. The first group included 65 rice (O. sativa L.) accessions introduced from seven countries, including five regions of
China. The second group included 58 US rice varieties released in the past 25 years. The third group consisted of 54 accessions of rice wild
relatives represented by ten different species. The number of alleles per SSR marker ranged from 4 to 32 with a mean of 16 alleles and the
polymorphism information content values ranged from 0.43 to 0.91 with a mean of 0.70. The variation in SSR alleles was a significant
contribution to the genetic discrimination of the 177 accessions within the three Oryza groups. Analysis of molecular variance identified deviation
from Hardy–Weinberg equilibrium. Principal coordinates analysis clearly separated the accessions into their respective three groups.
Neighbor-joining phylogenetic cluster reflects the ordination of each accession. Linkage disequilibrium (D′) averaged 0.75 in wild Oryza spp., and
about 0.5 in both US and international O. sativa accessions. Our results showed that LD among adjacent loci in both O. sativa and Oryza spp.
accessions is strong enough to be detecting marker-trait association via genome-wide scans.
049《Plant Molecular Biology》
Genetic and genomic approaches to develop rice germplasm for problem soils
Abdelbagi M. Ismail, Sigrid Heuer, Michael J. Thomson, Matthias Wissuwa
International Rice Research Institute, DAPO Box 7777, Metro Manila, Philippines
Abstract:Soils that contain toxic amounts of minerals or are deficient in essential plant nutrients are widespread globally and seriously constrain
rice production. New methods are necessary to incorporate the complex adaptive traits associated with tolerance of these abiotic stresses, while
simultaneously retaining the high yield potential of rice varieties when conditions are favorable. Significant progress in the genetic characterization
of stress response pathways and recent advances in genomics have provided powerful tools for in-depth dissection of tolerance mechanisms.
Additionally, tolerance of most of these abiotic stresses in rice is controlled by a few QTLs with large effects despite the intricacy of the numerous
traits involved. Genetic dissection of these QTLs and their incorporation into high-yielding varieties will significantly enhance and stabilize rice
productivity in these problem soils. Current efforts at IRRI and in rice breeding programs worldwide are seeking to explore diverse germplasm
collections and genetically dissect the causal mechanisms of tolerance to facilitate their use in breeding. This review focuses on salinity and P and
Zn deficiency as the major problems encountered in rice soils, and examines current understanding of the mechanisms involved and efforts toward
germplasm improvement.
050《Plant Molecular Biology》
A rice phenomics study—phenotype scoring and seed propagation of a T-DNA insertion-induced rice mutant population
Chyr-Guan Chern, Ming-Jen Fan, Su-May Yu, Ai-Ling Hour, Po-Chang Lu, Yao-Cheng Lin, Fu-Jin Wei, Sheng-Chung Huang, Shu Chen,
Ming-Hsing Lai, Ching-Shan Tseng, Hsing-Mu Yen, Woei-Shyuan Jwo, Chen-Chia Wu, Tung-Lung Yang, Lung-Sheng Li, Yih-Cheng Kuo,
Su-Mien Li, Charng-Pei Li, Chiu-Kai Wey, Arunee Trisiriroj, Hsing-Fang Lee, Yue-Ie C. Hsing
Taiwan Agricultural Research Institute, Wufeng, Taichung, 41301, Taiwan, China
Abstract:With the completion of the rice genome sequencing project, the next major challenge is the large-scale determination of gene function. As
an important crop and a model organism, rice provides major insights into gene functions important for crop growth or production. Phenomics
with detailed information about tagged populations provides a good tool for functional genomics analysis. By a T-DNA insertional mutagenesis
approach, we have generated a rice mutant population containing 55,000 promoter trap and gene activation or knockout lines. Approximately
20,000 of these lines have known integration sites. The T0 and T1 plants were grown in net “houses” for two cropping seasons each year since
2003, with the mutant phenotypes recorded. Detailed data describing growth and development of these plants, in 11 categories and 65
subcategories, over the entire four-month growing season are available in a searchable database, along with the genetic segregation information
and flanking sequence data. With the detailed data from more than 20,000 T1 lines and 12 plants per line, we estimated the mutation rates of the T1
population, as well the frequency of the dominant T0 mutants. The correlations among different mutation phenotypes are also calculated. Together,
the information about mutant lines, their integration sites, and the phenotypes make this collection, the Taiwan Rice Insertion Mutants (TRIM), a
good resource for rice phenomics study. Ten T2 seeds per line can be distributed to researchers upon request.
051《Plant Molecular Biology》
T-DNA tagged knockout mutation of rice OsGSK1 , an orthologue of Arabidopsis BIN2 , with enhanced tolerance to various abiotic stresses
Serry Koh, Sang-Choon Lee, Min-Kyung Kim, Jun Ho Koh, Sichul Lee, Gynheung An, Sunghwa Choe, Seong-Ryong Kim
Department of Life Science, Sogang University, Seoul, 121-742, Korea
Abstract:T-DNA-tagged rice plants were screened under cold- or salt-stress conditions to determine the genes involved in the molecular
mechanism for their abiotic-stress response. Line 0-165-65 was identified as a salt-responsive line. The gene responsible for this GUS-positive
phenotype was revealed by inverse PCR as OsGSK1 (O ryza s ativa g lycogen s ynthase k inase3-like gene 1), a member of the plant
GSK3/SHAGGY-like protein kinase genes and an orthologue of the Arabidopsis b rassinosteroid in sensitive 2 (BIN2), AtSK21. Northern blot
analysis showed that OsGSK1 was most highly detected in the developing panicles, suggesting that its expression is developmental stage specific.
Knockout (KO) mutants of OsGSK1 showed enhanced tolerance to cold, heat, salt, and drought stresses when compared with non-transgenic
segregants (NT). Overexpression of the full-length OsGSK1 led to a stunted growth phenotype similar to the one observed with the
gain-of-function BIN/AtSK21 mutant. This suggests that OsGSK1 might be a functional rice orthologue that serves as a negative regulator of
brassinosteroid (BR)-signaling. Therefore, we propose that stress-responsive OsGSK1 may have physiological roles in stress signal-transduction
pathways and floral developmental processes.
052《Molecular Genetics and Genomics》
Genomewide computational analysis of nitrate response elements in rice and Arabidopsis
Suman K. Das, Ravi Ramesh Pathak, Devapriya Choudhury, Nandula Raghuram
School of Biotechnology, Guru Gobind Singh Indraprastha University, Kashmiri Gate, Delhi, 110 006, India
Abstract:Nitrate response element (NRE) was originally reported to be comprised of an Ag/cTCA core sequence motif preceded by a 7-bp AT rich
region, based on promoter deletion analyses in nitrate and nitrite reductases from Arabidopsis thaliana and birch. In view of hundreds of new
nitrate responsive genes discovered recently, we sought to computationally verify whether the above motif indeed qualifies to be the cis-acting
NRE for all the responsive genes. We searched for the specific occurrence of at least two copies of the above motif in and around the nitrate
responsive genes and elsewhere in the Arabidopsis and rice (Oryza sativa) genomes, with respect to their positional, orientational and
strand-specific bias. This is the first comprehensive analysis of NREs for 625 nitrate responsive genes of Arabidopsis and their rice homologs,
representing dicots and monocots, respectively. We report that the above motifs are present almost randomly throughout these genomes and do not
reveal any specificity or bias towards nitrate responsive genes. This also seems to be true for smaller subsets of nitrate responsive genes in
Arabidopsis, such as the 21 early responsive genes, 261 and 90 genes for root-specific and shoot-specific response, respectively, and 25
housekeeping genes. This necessitates a fresh search for candidate sequences that qualify to be NREs in these and other plants.
053《Theoretical and Applied Genetics》
A quantitative trait locus for cold tolerance at the booting stage on rice chromosome 8
Makoto Kuroki, Koji Saito, Shuichi Matsuba, Narifumi Yokogami, Hiroyuki Shimizu, Ikuo Ando, Yutaka Sato
National Agricultural Research Center for Hokkaido Region, National Agriculture and Food Research Organization (NARO), 1 Hitsujigaoka,
Toyohira, Sapporo Hokkaido, 062-8555, Japan
Abstract:A quantitative trait locus (QTL) for cold tolerance at the booting stage of a cold-tolerant rice breeding line, Hokkai-PL9, was analyzed. A
total of 487 simple sequence repeat (SSR) markers distributed throughout the genome were used to survey for polymorphism between Hokkai-PL9
and a cold-sensitive breeding line, Hokkai287, and 54 markers were polymorphic. Single marker analysis revealed that markers on chromosome 8
are associated with cold tolerance. By interval mapping using an F 2 population between Hokkai-PL9 and Hokkai287, a QTL for cold tolerance was
detected on the short arm of chromosome 8. The QTL explains 26.6% of the phenotypic variance, and its additive effect is 11.4%. Substitution
mapping suggested that the QTL is located in a 193-kb interval between SSR markers RM5647 and PLA61. We tentatively designated the QTL as
qCTB8 (quantitative trait locus for cold tolerance at the booting stage on chromosome 8).
054《Theoretical and Applied Genetics》
A marker-assisted backcross approach for developing submergence-tolerant rice cultivars
C. N. Neeraja, R. Maghirang-Rodriguez, A. Pamplona, S. Heuer, B. C. Y. Collard, E. M. Septiningsih, G. Vergara, D. Sanchez, K. Xu, A. M. Ismail,
D. J. Mackill
International Rice Research Institute, DAPO Box 7777, Metro Manila, Philippines
Abstract:Submergence stress regularly affects 15 million hectares or more of rainfed lowland rice areas in South and Southeast Asia. A major QTL
on chromosome 9, Sub1, has provided the opportunity to apply marker assisted backcrossing (MAB) to develop submergence tolerant versions of
rice cultivars that are widely grown in the region. In the present study, molecular markers that were tightly linked with Sub1, flanking Sub1, and
unlinked to Sub1 were used to apply foreground, recombinant, and background selection, respectively, in backcrosses between a
submergence-tolerant donor and the widely grown recurrent parent Swarna. By the BC2F2 generation a submergence tolerant plant was identified
that possessed Swarna type simple sequence repeat (SSR) alleles on all fragments analyzed except the tip segment of rice chromosome 9 that
possessed the Sub1 locus. A BC3F2 double recombinant plant was identified that was homozygous for all Swarna type alleles except for an
approximately 2.3–3.4 Mb region surrounding the Sub1 locus. The results showed that the mega variety Swarna could be efficiently converted to a
submergence tolerant variety in three backcross generations, involving a time of two to three years. Polymorphic markers for foreground and
recombinant selection were identified for four other mega varieties to develop a wider range of submergence tolerant varieties to meet the needs of
farmers in the flood-prone regions. This approach demonstrates the effective use of marker assisted selection for a major QTL in a molecular
breeding program
055《Euphytica》
Comparative genetic analysis and molecular mapping of fertility restoration genes for WA, Dissi, and Gambiaca cytoplasmic male sterility systems
in rice
Majid Sattari, Arumugam Kathiresan, Glenn B. Gregorio, Sant S. Virmani
Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute (IRRI), DAPO Box 7777, Metro Manila, Philippines
Abstract:The genetic relationship among three cytoplasmic male sterility (CMS) systems, consisting of WA, Dissi, and Gambiaca, was studied.
The results showed that the maintainers of one CMS system can also maintain sterility in other cytoplasmic backgrounds. The F 1 plants derived
from crosses involving A and R lines of the respective cytoplasm and their cross-combination with other CMS systems showed similar pollen and
spikelet fertility values, indicating that similar biological processes govern fertility restoration in these three CMS systems. The results from an
inheritance study showed that the pollen fertility restoration in all three CMS systems was governed by two independent and dominant genes with
classical duplicate gene action. Three F2 populations, generated from the crosses between the parents of good-performing rice hybrids, that possess
WA, Dissi, and Gambiaca CMS cytoplasm, were used to map the Rf genes. For the WA-CMS system, Rf3 was located at a distance of 2.8 cM from
RM490 on chromosome 1 and Rf4 was located at 1.6 cM from RM1108 on chromosome 10. For the Dissi-CMS system, Rf3 was located on
chromosome 1 at 1.9 cM from RM7466 and Rf4 on chromosome 10 was located at 2.3 cM from RM6100. The effect of Rf3 on pollen fertility
appeared to be stronger than the effect of Rf4. In the Gambiaca-CMS system, only one major locus was mapped on chromosome 1 at 2.1 cM from
RM576. These studies have led to the development of marker-assisted selection (MAS) for selecting putative restorer lines, new approaches to
alloplasmic line breeding, and the transfer of Rf genes into adapted cultivars through a backcrossing program in an active hybrid rice breeding
program.
056《Plant Molecular Biology》
Elicitor induced activation of the methylerythritol phosphate pathway toward phytoalexins biosynthesis in rice
Atsushi Okada, Takafumi Shimizu, Kazunori Okada, Tomohisa Kuzuyama, Jinichiro Koga, Naoto Shibuya, Hideaki Nojiri, Hisakazu Yamane
Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
Abstract:Diterpenoid phytoalexins such as momilactones and phytocassanes are produced via geranylgeranyl diphosphate in suspension-cultured
rice cells after treatment with a chitin elicitor. We have previously shown that the production of diterpene hydrocarbons leading to phytoalexins
and the expression of related biosynthetic genes are activated in suspension-cultured rice cells upon elicitor treatment. To better understand the
elicitor-induced activation of phytoalexin biosynthesis, we conducted microarray analysis using suspension-cultured rice cells collected at various
times after treatment with chitin elicitor. Hierarchical cluster analysis revealed two types of early-induced expression (EIE-1, EIE-2) nodes and a
late-induced expression (LIE) node that includes genes involved in phytoalexins biosynthesis. The LIE node contains genes that may be
responsible for the methylerythritol phosphate (MEP) pathway, a plastidic biosynthetic pathway for isopentenyl diphosphate, an early precursor of
phytoalexins. The elicitor-induced expression of these putative MEP pathway genes was confirmed by quantitative reverse-transcription PCR.
1-Deoxy-d-xylulose 5-phosphate synthase (DXS), 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR), and 4-(cytidine
5′-diphospho)-2-C-methyl-d-erythritol synthase (CMS), which catalyze the first three committed steps in the MEP pathway, were further shown to
have enzymatic activities that complement the growth of E. coli mutants disrupted in the corresponding genes. Application of ketoclomazone and
fosmidomycin, inhibitors of DXS and DXR, respectively, repressed the accumulation of diterpene-type phytoalexins in suspension cells treated
with chitin elicitor. These results suggest that activation of the MEP pathway is required to supply sufficient terpenoid precursors for the
production of phytoalexins in infected rice plants.
057《Euphytica》
Local coexpression domains in the genome of rice show no microsynteny with Arabidopsis domains
Xin-Ying Ren, Willem J. Stiekema, Jan-Peter Nap
Applied Bioinformatics, Plant Research International, Plant Sciences Group, Wageningen University and Research Centre, P.O. Box 16, 6700 AA
Wageningen, The Netherlands
Abstract:Chromosomal coexpression domains are found in a number of different genomes under various developmental conditions. The size of
these domains and the number of genes they contain vary. Here, we define local coexpression domains as adjacent genes where all possible
pair-wise correlations of expression data are higher than 0.7. In rice, such local coexpression domains range from predominantly two genes, up to 4,
and make up ∼5% of the genomic neighboring genes, when examining different expression platforms from the public domain. The genes in local
coexpression domains do not fall in the same ontology category significantly more than neighboring genes that are not coexpressed. Duplication,
orientation or the distance between the genes does not solely explain coexpression. The regulation of coexpression is therefore thought to be
regulated at the level of chromatin structure. The characteristics of the local coexpression domains in rice are strikingly similar to such domains in
the Arabidopsis genome. Yet, no microsynteny between local coexpression domains in Arabidopsis and rice could be identified. Although the rice
genome is not yet as extensively annotated as the Arabidopsis genome, the lack of conservation of local coexpression domains may indicate that
such domains have not played a major role in the evolution of genome structure or in genome conservation.
058《Molecular Genetics and Genomics》
Expression analysis of calcium-dependent protein kinase gene family during reproductive development and abiotic stress conditions in rice ( Oryza
sativa L. ssp. indica )
Swatismita Ray, Pinky Agarwal, Rita Arora, Sanjay Kapoor, Akhilesh K. Tyagi
Interdisciplinary Centre for Plant Genomics and Department of Plant Molecular Biology, University of Delhi South Campus, Benito Juarez Road,
New Delhi, 110021, India
Abstract:Calcium-dependent protein kinases (CDPKs) are important sensors of Ca+2 flux in plants, which control plant development and responses
by regulating downstream components of calcium signaling pathways. Availability of the whole genome sequence and microarray platform allows
investigation of genome-wide organization and expression profile of CDPK genes in rice with a view to ultimately define their function in plant
systems. Genome-wide analysis led to identification of 31 CDPK genes in rice after a thorough annotation exercise based upon HMM profiles.
Twenty-nine already identified CDPK genes were verified and two new members were added to the CDPK gene family of rice. Relative
expression of all these genes has been analyzed by using Affymetrix rice genome array™ during three vegetative stages, six stages of panicle
(P1–P6) and five stages of seed (S1–S5) development along with three abiotic stress conditions, viz. cold, salt and desiccation, given to seedling.
Thirty-one CDPK genes were found to express in at least one of the experimental stages studied. Of these, transcripts for twenty three genes
accumulated differentially during reproductive developmental stages; nine of them were preferentially up-regulated only in panicle, five were
up-regulated in stages of panicles as well as seed development, whereas, expression of one gene was found to be specific to the S1 stage of seed
development. Eight genes were found to be down-regulated during the panicle and seed developmental stages. Six CDPK genes were found to be
induced while the expression of one gene was down-regulated under stress conditions. The differential expression of CDPK genes during
reproductive development and stress is suggestive of their involvement in the underlying signal transduction pathways. Furthermore, up-regulation
of common genes both during reproductive development as well as stress responses is indicative of common element between reproduction and
stress.
059《Journal of Integrative Plant Biology》
The WRKY Gene Family in Rice (Oryza sativa)
Christian A. Ross, Yue Liu and Qingxi J. Shen
Bioinformatics Core, School of Life Sciences, University of Nevada, Las Vegas, Nevada 89154, USA
Abstract:WRKY genes encode transcription factors that are involved in the regulation of various biological processes. These zinc-finger proteins,
especially those members mediating stress responses, are uniquely expanded in plants. To facilitate the study of the evolutionary history and
functions of this supergene family, we performed an exhaustive search for WRKY genes using HMMER and a Hidden Markov Model that was
specifically trained for rice. This work resulted in a comprehensive list of WRKY gene models in Oryza sativa L. ssp. indica and L. ssp. japonica.
Mapping of these genes to individual chromosomes facilitated elimination of the redundant, leading to the identification of 98 WRKY genes in
japonica and 102 in indica rice. These genes were further categorized according to the number and structure of their zinc-finger domains. Based on
a phylogenetic tree of the conserved WRKY domains and the graphic display of WRKY loci on corresponding indica and japonica chromosomes,
we identified possible WRKY gene duplications within, and losses between the two closely related rice subspecies. Also reviewed are the roles of
WRKY genes in disease resistance and responses to salicylic acid and jasmonic acid, seed development and germination mediated by gibberellins,
other developmental processes including senescence, and responses to abiotic stresses and abscisic acid in rice and other plants. The signaling
pathways mediating WRKY gene expression are also discussed.
060《Plant Science》
Molecular characterization of a novel isoform of rice (Oryza sativa L.) glycine rich-RNA binding protein and evidence for its involvement in high
temperature stress response
Chandan Sahi, Manu Agarwal, Amanjot Singh and Anil Grover
Department of Plant Molecular Biology, University of Delhi South Campus, Benito Juarez Road, Dhaula Kuan, New Delhi 110021, India
Abstract:A novel full-length cDNA encoding for glycine rich (GR)-RNA binding protein (RBP) (Osgr-rbp4) is isolated from rice heat shock cDNA
library. Amino acid sequence of the deduced protein reveals existence of RNA recognition motif (RRM) comprising of highly conserved RNA
binding RNPI and RNPII domains at the N-terminus. C-terminus of this protein is rich in arginine and glycine residues. Blast search analysis on
rice genome sequence database shows that GR-RBP protein family is constituted of multiple members with high level of amino acid conservation
in RNA recognition motif and glycine domain regions. Similar analysis across wider biological systems from NCBI database indicated that rice
GR-RBP4 has homologs in different living genera. Osgr-rbp4 transcript in rice seedlings is constitutively expressed as well as regulated by
different abiotic stresses including high temperature stress. Ectopic over-expression of Osgr-rbp4 cDNA imparts high temperature stress tolerance
to wild type yeast cells. It is shown that OsGR-RBP4 in rice leaf cells and its immunologically homologous protein in tobacco BY2 protoplasts are
nuclear proteins. Upon heat shock, bulk of these proteins appears to be localized in the cytoplasm. We suggest that OsGR-RBP4 probably bind and
stabilize the stress-inducible transcripts under HS conditions.
061《Plant Molecular Biology》
Genome-wide identification of C2H2 zinc-finger gene family in rice and their phylogeny and expression analysis
Pinky Agarwal, Rita Arora, Swatismita Ray, Ashok K. Singh, Vijay P. Singh, Hiroshi Takatsuji, Sanjay Kapoor, Akhilesh K. Tyagi
Interdisciplinary Centre for Plant Genomics and Department of Plant Molecular Biology, University of Delhi South Campus, Benito Juarez Road,
New Delhi, 110021, India
Abstract:Transcription factors regulate gene expression in response to various external and internal cues by activating or suppressing downstream
genes in a pathway. In this study, we provide a complete overview of the genes encoding C 2H2 zinc-finger transcription factors in rice, describing
the gene structure, gene expression, genome localization, and phylogenetic relationship of each member. The genome of Oryza sativa codes for
189 C2H2 zinc-finger transcription factors, which possess two main types of zinc-fingers (named C and Q). The Q-type zinc fingers contain a
conserved motif, QALGGH, and are plant specific, whereas C type zinc fingers are found in other organisms as well. A genome-wide microarray
based gene expression analysis involving 14 stages of vegetative and reproductive development along with 3 stress conditions has revealed that
C2H2 gene family in indica rice could be involved during all the stages of reproductive development from panicle initiation till seed maturation. A
total of 39 genes are up-regulated more than 2-fold, in comparison to vegetative stages, during reproductive development of rice, out of which 18
are specific to panicle development and 12 genes are seed-specific. Twenty-six genes have been found to be up-regulated during three abiotic
stresses and of these, 14 genes express specifically during the stress conditions analyzed while 12 are also up-regulated during reproductive
development, suggesting that some components of the stress response pathways are also involved in reproduction.
062《Gene》
Characterization of a multifunctional inositol phosphate kinase from rice and barley belonging to the ATP-grasp superfamily
Lone Josefsen, Lisbeth Bohn, Mikael Blom Sørensen and Søren K. Rasmussen
Department of Agricultural Sciences, Faculty of Life Science, University of Copenhagen, DK-1871 Frederiksberg C, Denmark
Abstract:OsIpk and HvIpk, inositol phosphate kinases, were cloned from rice (Oryza sativa L. var. indica, IR64) and barley (Hordeum vulgare)
respectively. Sequence alignment showed that they belong to the ATP-grasp family, which includes inositol 1,3,4-trisphosphate 5/6-kinase from
humans and Arabidopsis. Residues that are binding sites for ATP and coordinate magnesium in absence or presence of inositol phosphate are
conserved and in total 23 residues are invariant among the twelve aligned inositol phosphate kinases. The genes were heterologously expressed in
Escherichia coli and kinase activity assays with 17 different isomers of inositol mono-/di-/tri-/tetra-/pentaphosphate as well as phytate were
performed. The strongest activity for both kinases was observed with Ins(3,4,5,6)P4, which candidates as the primary substrate for these kinases in
plants. Several species-specific differences between the two recombinant Ipks were observed. Rice OsIpk showed detectable kinase activity
towards eight different substrates, whereas barley HvIpk showed kinase activity with all the substrates including inositol mono- and bisphosphates.
HvIpk showed 3-kinase activity towards the Ins(1,4,5)P 3 substrate and it also interconverted the two substrates Ins(1,3,4,5)P 4 and Ins(1,3,4,6)P4 by
isomerase activity, which was not observed for the rice homologue. Both OsIpk and HvIpk had no detectable 2-kinase activity. Furthermore, the
two Ipks showed phosphatase activity towards several inositol phosphates. Expression analysis by RT-PCR demonstrated that the Ipk gene was
equally expressed in different tissues and developmental stages. Taken together, these results show that the Ipk kinase plays a significant role in the
inositol phosphate interacting network in plants.
063《Plant Molecular Biology》
Mutations in the rice liguleless gene result in a complete loss of the auricle, ligule, and laminar joint
Jinwon Lee, Jong-Jin Park, Song Lim Kim, Jieun Yim, Gynheung An
National Research Laboratory of Plant Functional Genomics, Division of Molecular and Life Sciences, Pohang University of Science and
Technology (POSTECH), Hyo-ja dong, Pohang, Kyungbuk, 790-784, Korea
Abstract:The area between the upper part of the leaf sheath and the basal portion of the leaf blade contains several specialized organs, such as the
laminar joint, auricle and ligule. Here we report the identification of T-DNA insertional mutant lines that lack all of these organs. The gene
knocked out in the mutant lines encodes a protein that contains a SBP (SQUAMOSA promoter Binding Protein)-domain and is highly homologous
to the maize LIGULELESS1 (LG1) gene. At the amino acid sequence level, the OsLG1 protein is 69% identical to maize LG1 and 78% identical
to barley LG1. We named the rice gene OsLIGULELESS1 (OsLG1). Transient expression of an OsLG1:RFP (Red Fluorescent Protein) fusion
protein indicated that the protein is localized to the nucleus. Transgenic plants harboring the OsLG1 promoter:GUS (β-glucuronidase) reporter
gene construct display preferential expression in developing laminar joint regions and meristemic regions. The gene is also weakly expressed in
the ligule, auricles, and leaf sheaths at the basal region. These results indicate that OsLG1 is a transcriptional factor that plays an important role in
building the laminar joint between leaf blade and leaf sheath boundary, thereby controlling ligule and auricle development.
064《Plant Cell Reports》
Regeneration of transgenic plants from two indica rice ( Oryza sativa L.) cultivars using shoot apex explants
S. Arockiasamy, S. Ignacimuthu
Entomology Research Institute, Loyola College, Chennai, 600034, India
Abstract:We have established a reproducible procedure for transformation of shoot apices and regeneration of transgenic plants for two indica rice
cultivars, white ponni (WP) and Pusa Basmathi 1 (PB 1). Four-day-old shoot apex explants were transformed by cocultivation with Agrobacterium
tumefaciens strain EHA 101 harbouring a binary plasmid pRIT1. The vector contained an improved hygromycin phosphotransferase (hpt) gene for
hygromycin resistance driven by actin 1 promoter and the reporter gene β-glucuronidase intron (INT-GUS) controlled by CaMV 35S promoter.
Rice shoots were induced on media containing 0.1 mg/l napthalene acetic acid (NAA), 1.0 mg/l kinetin (kn), 1.0 mg/l N6-benzyleaminopurin
(BAP), 300 mg/l casaminoacid, 500 mg/l proline, 50 mg/l hygromycin and 500 mg/l cefotaxime. Transgenic plants were raised in pots and seeds
were collected. Histochemical and polymerase chain reaction (PCR) analyses of field established transgenic rice plants and their offsprings
confirmed the presence of GUS gene. Integration of T-DNA into the genome of putative transgenics was further confirmed by southern analysis.
The transformation efficiency of WP was found to be ranging from 5.6 to 6.2% whereas in the case of PB1, it was from 7 to 8%. Progeny analysis
of these plants showed a pattern of classical Mendelian inheritance for both hpt and GUS gene.
065《Theoretical and Applied Genetics》
The identification of candidate rice genes that confer resistance to the brown planthopper ( Nilaparvata lugens ) through representational difference
analysis
Dong-Soo Park, Sang-Kyu Lee, Jong-Hee Lee, Min-Young Song, Song-Yi Song, Do-Yeon Kwak, Un-Sang Yeo, Nam-Soo Jeon, Soo-Kwon Park,
Gihwan Yi, You-Chun Song, Min-Hee Nam, Yeon-Chung Ku, Jong-Seong Jeon
Yeongnam Agricultural Research Institute, Milyang, 627-803, South Korea
Abstract:The development of rice varieties (Oryza sativa L.) that are resistant to the brown planthopper (BPH; Nilaparvata lugens Stål) is an
important objective in current breeding programs. In this study, we generated 132 BC 5F5 near-isogenic rice lines (NILs) by five backcrosses of
Samgangbyeo, a BPH resistant indica variety carrying the Bph1 locus, with Nagdongbyeo, a BPH susceptible japonica variety. To identify genes
that confer BPH resistance, we employed representational difference analysis (RDA) to detect transcripts that were exclusively expressed in one of
our BPH resistant NIL, SNBC61, during insect feeding. The chromosomal mapping of the RDA clones that we subsequently isolated revealed that
they are located in close proximity either to known quantitative trait loci or to an introgressed SSR marker from the BPH resistant donor parent
Samgangbyeo. Genomic DNA gel-blot analysis further revealed that loci of all RDA clones in SNBC61 correspond to the alleles of Samgangbyeo.
Most of the RDA clones were found to be exclusively expressed in SNBC61 and could be assigned to functional groups involved in plant defense.
These RDA clones therefore represent candidate defense genes for BPH resistance.
066《Molecular Breeding》
Mapping of quantitative trait loci for basmati quality traits in rice ( Oryza sativa L.)
Yellari Amarawathi, Rakesh Singh, Ashok K. Singh, Vijai P. Singh, Trilochan Mohapatra, Tilak R. Sharma, Nagendra K. Singh
Rice Genome Laboratory, National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute, Lal Bahadur Shastri Building,
Pusa Campus, New Delhi, Delhi, 110012, India
Abstract:Traditional basmati rice varieties are very low yielding due to their poor harvest index, tendency to lodging and increasing susceptibility
to foliar diseases; hence there is a need to develop new varieties combining the grain quality attributes of basmati with high yield potential to fill
the demand gap. Genetic control of basmati grain and cooking quality traits is quite complex, but breeding work can be greatly facilitated by use of
molecular markers tightly linked to these traits. A set of 209 recombinant inbred lines (RILs) developed from a cross between basmati quality
variety Pusa 1121 and a contrasting quality breeding line Pusa 1342, were used to map the quantitative trait loci (QTLs) for seven important
quality traits namely grain length (GL), grain breadth (GB), grain length to breadth ratio (LBR), cooked kernel elongation ratio (ELR), amylose
content (AC), alkali spreading value (ASV) and aroma. A framework molecular linkage map was constructed using 110 polymorphic simple
sequence repeat (SSR) markers distributed over the 12 rice chromosomes. A number of QTLs, including three for GL, two for GB, two for LBR,
three for aroma and one each for ELR, AC and ASV were mapped on seven different chromosomes. While location of majority of these QTLs was
consistent with the previous reports, one QTL for GL on chromosomes 1, and one QTL each for ELR and aroma on chromosomes 11 and 3,
respectively, are being reported here for the first time. Contrary to the earlier reports of monogenic recessive inheritance, the aroma in Pusa 1121 is
controlled by at least three genes located on chromosomes 3, 4 and 8, and similar to the reported association of badh2 gene with aroma QTL on
chromosome 8, we identified location of badh1 gene in the aroma QTL interval on chromosome 4. A discontinuous 5 + 3 bp deletion in the seventh
exon of badh2 gene, though present in all the RILs with high aroma, was not sufficient to impart this trait to the rice grains as many of the RILs
possessing this deletion showed only mild or no aroma expression.
067《Plant Cell Reports》
Transgenic tobacco plants overexpressing the heterologous lea gene Rab16A from rice during high salt and water deficit display enhanced
tolerance to salinity stress
Aryadeep RoyChoudhury, Chaitali Roy, Dibyendu N. Sengupta
Department of Botany, Bose Institute, 93/1, Acharya Prafulla Chandra Road, Kolkata, 700009, West Bengal, India
Abstract:The full length Rab16A, from the indica rice Pokkali, was introduced into tobacco by Agrobacterium-mediated transformation. The
transgene was stably integrated into the genome and they originated from different lines of integration. Expression of Rab16A transcript driven by
its own promoter (stress inducible) in T2 progenies, only when triggered by salinity/ABA/PEG (Polyethylene glycol)-mediated dehydration, but
not at the constitutive level, led to the stress-induced accumulation of RAB16A protein in the leaves of transgenic plants. The selected independent
transgenic lines showed normal growth, morphology and seed production as the WT plants without any yield penalty under stress conditions. They
exhibited significantly increased tolerance to salinity, sustained growth rates under stress conditions; with concomitant increased osmolyte
production like reducing sugars, proline and higher polyamines. They also showed delayed development of damage symptoms with better
antioxidative machinery and more favorable mineral balance, as reflected by reduced H 2O2 levels and lipid peroxidation, lesser chlorophyll loss as
well as lesser accumulation of Na+ and greater accumulation of K+ in 200 mM NaCl. These findings establish the potential role of Rab16A gene in
conferring salt tolerance without affecting growth and yield, as well as pointing to the fact that the upstream region of Rab16A behaves as an
efficient stress-inducible promoter. Our result also suggests the considerable potential of Group 2 lea genes as molecular tools for genetic
engineering of plants towards stress tolerance.
068《Plant Biotechnology Journal》
Emerging trends in the functional genomics of the abiotic stress response in crop plants
Shubha Vij, Akhilesh K. Tyagi
Interdisciplinary Centre for Plant Genomics and Department of Plant Molecular Biology, University of Delhi South Campus, New Delhi 110 021,
India
Abstract:Plants are exposed to different abiotic stresses, such as water deficit, high temperature, salinity, cold, heavy metals and mechanical
wounding, under field conditions. It is estimated that such stress conditions can potentially reduce the yield of crop plants by more than 50%.
Investigations of the physiological, biochemical and molecular aspects of stress tolerance have been conducted to unravel the intrinsic mechanisms
developed during evolution to mitigate against stress by plants. Before the advent of the genomics era, researchers primarily used a gene-by-gene
approach to decipher the function of the genes involved in the abiotic stress response. However, abiotic stress tolerance is a complex trait and,
although large numbers of genes have been identified to be involved in the abiotic stress response, there remain large gaps in our understanding of
the trait. The availability of the genome sequences of certain important plant species has enabled the use of strategies, such as genome-wide
expression profiling, to identify the genes associated with the stress response, followed by the verification of gene function by the analysis of
mutants and transgenics. Certain components of both abscisic acid-dependent and -independent cascades involved in the stress response have
already been identified. Information originating from the genome-wide analysis of abiotic stress tolerance will help to provide an insight into the
stress-responsive network(s), and may allow the modification of this network to reduce the loss caused by stress and to increase agricultural
productivity.
069《Transgenic Research》
Over expression of cytosolic copper/zinc superoxide dismutase from a mangrove plant Avicennia marina in indica Rice var Pusa Basmati-1 confers
abiotic stress tolerance
S. R. Prashanth, V. Sadhasivam, Ajay Parida
M.S. Swaminathan Research Foundation, 3rd cross street, Taramani Institutional area, Chennai, 600113, India
Abstract:Antioxidant enzymes play an important role in conferring abiotic stress tolerance. Superoxide dismutase (SOD) is the first enzyme in the
enzymatic antioxidative pathway. Halophytic plants like mangroves have been reported to have a high level of SOD activity, which plays a major
role in defending the mangrove species against severe abiotic stresses. We had previously reported the isolation of Sod1, a cDNA encoding a
cytosolic copper zinc superoxide dismutase from the mangrove plant Avicennia marina and its mRNA expression pattern during various oxidative
and abiotic stresses. The present study is an extension of the previous study in further characterizing the Sod1 cDNA by transforming it into rice
and analysing the transgenic plants for abiotic stress tolerance. Southern hybridization of A. marina genomic DNA using Sod1, revealed that this
gene in A. marina genome is present as a single copy. The cDNA was cloned into a binary vector (pCAMBIA 1300) and transformed into indica
rice var Pusa Basmati-1. Southern hybridization analysis of transgenic rice plants revealed stable integration of the Sod1 transgene in the rice
genome. The mRNA transcript of Sod1 was detected by Northern hybridisation in the transgenic rice plants. SOD isozyme assay of the transgenic
rice plants revealed the stable expression of the transgenic Sod1 protein. The transgenic plants were more tolerant to methyl viologen mediated
oxidative stress in comparison to the untransformed control plants. The transgenic plants also withstood salinity stress of 150 mM of NaCl for a
period of eight days while the untransformed control plants wilted at the end of the stress treatment in hydroponics. Pot grown transgenic plants
could also tolerate salinity stress better than the untransformed control plants, when irrigated with saline water. The transgenic plants also revealed
better tolerance to drought stress in comparison to untransformed control plants.
070《Journal of Biosciences》
Complexity of rice Hsp100 gene family: lessons from rice genome sequence data
Gaurav Batra, Vineeta Singh Chauhan, Amanjot Singh, Neelam K Sarkar, Anil Grover
Department of Plant Molecular Biology, University of Delhi South Campus, New Delhi, 110 021, India
Abstract:Elucidation of genome sequence provides an excellent platform to understand detailed complexity of the various gene families. Hsp100 is
an important family of chaperones in diverse living systems. There are eight putative gene loci encoding for Hsp100 proteins in Arabidopsis
genome. In rice, two full-length Hsp100 cDNAs have been isolated and sequenced so far. Analysis of rice genomic sequence by in silico approach
showed that two isolated rice Hsp100 cDNAs correspond to Os05g44340 and Os02g32520 genes in the rice genome database. There appears to be
three additional proteins (encoded by Os03g31300, Os04g32560 and Os04g33210 gene loci) that are variably homologous to Os05g44340 and
Os02g32520 throughout the entire amino acid sequence. The above five rice Hsp100 genes show significant similarities in the signature sequences
known to be conserved among Hsp100 proteins. While Os05g44340 encodes cytoplasmic Hsp100 protein, those encoded by the other four genes
are predicted to have chloroplast transit peptides.
071《Euphytica》
Molecular characterization of the recombinant inbred line population derived from a japonica-indica rice cross
Guangjie Liu, John L. Bernhardt, Melissa H. Jia, Yeshi A. Wamishe, Yulin Jia
Rice Research and Extension Center, University of Arkansas, 2900 Hwy 130E, Stuttgart, AR 72160, USA
Abstract:Recombinant inbred line (RIL) populations of rice are useful genetic sources for map-based cloning of agronomically important genes.
Zhe733 is a high-yielding indica cultivar from China conferring resistance to rice blast (RB), rice water weevil (RWW) and straighthead; whereas
Kaybonnet low-phytic acid 1-1 (KBNTlpa) is a mutant of a tropical japonica cultivar from the US containing low-phytic acid with average yield,
and is susceptible to some RB races, RWW, and straighthead. A 355 RIL F10−11 population derived from the cross of KBNTlpa × Zhe733 was
recently released. Simple sequence repeat (SSR) markers were used to evaluate 269 RILs of this population. A total of 107 polymorphic markers
were mapped on all rice chromosomes representing a total of 1,016.3 cM of genetic distance. Two hundred and thirty-five KBNTlpa × Zhe733
RILs (KZRILs) were clustered into seven groups based on allele frequencies of SSR markers. Twenty-three markers (21.1%) on chromosomes 3, 6,
7, 9, and 11 were found to favor Zhe733 (χ 2 = 16.8−189.7 and P < 0.01) and five markers (4.6%) on chromosome 1 and 6 were found to favor
KBNTlpa (χ 2 = 18.5−46.6 and P < 0.01). Marker segregations were observed to be normal for both parents except 26 (10.2%) KZRILs were
found to skew toward Zhe733 (χ 2 > 15.7 and P < 0.01). Furthermore, the average frequencies of heterozygosity and non-parental alleles per
KZRIL were 1.3% (0.0−38.9%) and 0.4% (0.0−15.0%), respectively. Thirteen heterozygous KZRILs were found at more than five markers loci
and nine KZRILs were found with more than five non-parental alleles representing 5.1 and 3.5% of 255 KZRILs. Overall, this KZRIL population
is a good population with relatively low frequencies of heterozygosity and non-parental alleles, and with relatively low percentages of skewed
markers and skewed KZRILs. The profiles of these SSR markers should facilitate molecular tagging critical genes controlling yield, RB, RWW,
and straighthead resistance.
072《Theoretical and Applied Genetics》
Leveraging the rice genome sequence for monocot comparative and translational genomics
H. C. Lohithaswa, F. A. Feltus, H. P. Singh, C. D. Bacon, C. D. Bailey, A. H. Paterson
Plant Genome Mapping Laboratory, University of Georgia, 111 Riverbend Road, Athens, GA 30602, USA
Abstract:Common genome anchor points across many taxa greatly facilitate translational and comparative genomics and will improve our
understanding of the Tree of Life. To add to the repertoire of genomic tools applicable to the study of monocotyledonous plants in general, we
aligned Allium and Musa ESTs to Oryza BAC sequences and identified candidate Allium-Oryza and Musa-Oryza conserved intron-scanning
primers (CISPs). A random sampling of 96 CISP primer pairs, representing loci from 11 of the 12 chromosomes in rice, were tested on seven
members of the order Poales and on representatives of the Arecales, Asparagales, and Zingiberales monocot orders. The single-copy amplification
success rates of Allium (31.3%), Cynodon (31.4%), Hordeum (30.2%), Musa (37.5%), Oryza (61.5%), Pennisetum (33.3%), Sorghum (47.9%),
Zea (33.3%), Triticum (30.2%), and representatives of the palm family (32.3%) suggest that subsets of these primers will provide DNA markers
suitable for comparative and translational genomics in orphan crops, as well as for applications in conservation biology, ecology, invasion biology,
population biology, systematic biology, and related fields. Electronic supplementary material The online version of this article
(doi:10.1007/s00122-007-0559-4) contains supplementary material, which is available to authorized users.
073《Plant Cell Reports》
Agrobacterium -mediated transformation of a low glutelin mutant of ‘Koshihikari’ rice variety using the mutated-acetolactate synthase gene
derived from rice genome as a selectable marker
Yuhya Wakasa, Kenjirou Ozawa, Fumio Takaiwa
Transgenic Crop Research and Development Center, National Institute of Agrobiological Sciences, Kannondai 3-1-3, Tsukuba Ibaraki, 305-8602,
Japan
Abstract:We have developed an efficient rice transformation system that uses only rice genome-derived components. The transgenic ‘Koshihikari’
rice, low-glutelin mutant a123, is capable of accumulating large amounts of bioactive peptides in the endosperm. Agrobacterium-mediated
transformation using the mutated-acetolactate synthase (mALS) gene expressed under the control of the callus-specific promoter (CSP) as a
selectable marker was used to introduce GFP and an anti-hypertensive hexapeptide into ‘Koshihikari’ a123. The CSP:mALS gene cassette confers
pyrimidinyl carboxy herbicide resistance to transgenic rice callus, but is not expressed in regenerated plants. Transformation efficiency of
transgenic rice line a123 was improved from about 10% to about 30% by modifying callus induction, callus selection and regeneration media
conventionally used for rice tissue culture.