Turkish Journal of Biology
Turk J Biol
(2016) 40: 554-560
© TÜBİTAK
doi:10.3906/biy-1503-30
http://journals.tubitak.gov.tr/biology/
Research Article
Molecular cytogenetic identification of a novel hexaploid wheat–Thinopyrum intermedium partial amphiploid with high protein content
1
1
2
1
2,
Mariyana GEORGIEVA , Klaudia KRUPPA , Nedyalka TYANKOVA , Márta MOLNÁR-LÁNG *
Department of Molecular Genetics, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Sofia, Bulgaria
2
Agricultural Institute, Centre for Agricultural Research, Hungarian Academy of Sciences, Martonvásár, Hungary
Received: 10.03.2015
Accepted/Published Online: 09.06.2015
Final Version: 18.05.2016
Abstract: Genomic in situ hybridization and fluorescence in situ hybridization were used to define the cytogenetic constitution of the
intergeneric introgression line 29(1-57) obtained from the wheat × Thinopyrum intermedium (wheatgrass) cross. This line is a highprotein line resistant to leaf rust, yellow rust, and powdery mildew under field conditions and has 42 chromosomes. GISH analysis using
a total genomic DNA probe from Thinopyrum intermedium resulted in the identification of 14 chromosomes of wheatgrass origin. The
sequential GISH patterns obtained using St-genomic and J-genomic DNA as probes and ABD-genomic DNA from wheat as a blocker
led to a better determination of the genomic type of wheatgrass chromosomes. Rearrangements involving Thinopyrum chromosomes
and the A, B, and D genomes of wheat were not observed. FISH using the repetitive DNA probes pSc119.2, Afa-family, and pTa71
allowed for identification of all the wheat chromosomes present.
Key words: FISH, GISH, wheat–Thinopyrum intermedium introgression line
1. Introduction
The use of alien germplasm has increased intensively
in cereal breeding programs for wheat improvement
(Whitford et al., 2013). The alien species involved in
the development of new amphiploids and wheat–alien
chromosome addition, substitution, and translocation
lines include the genera Aegilops (Schneider et al., 2005;
Kumar et al., 2010; Farkas et al., 2014), Thinopyrum
(Sepsi et al., 2008; Georgieva et al., 2011b; Zeng et al.,
2013), Dasypyrum (Chen et al., 1995; Yang et al., 2005),
Secale (Tang et al., 2008), Hordeum (Munns et al., 2011;
Molnár-Láng et al., 2014), Psathyrostachys (Wang et al.,
2011), etc. Despite major progress in plant production, the
development of salt- and drought-tolerant crops remains
elusive in the case of every important crop (Priya et al.,
2015). Crosses between Thinopyrum spp. and Triticum
aestivum L. contribute to disease resistance and tolerance
for soil acidity, drought, salinity, and waterlogging in
wheat.
The diversity of hybrids can also genetically enrich
the wheatgrass germplasm used for pasture, hay, and soil
erosion control (Turner et al., 2013). Wheat–wheatgrass
lines produced via crosses with Th. intermedium,
Th. ponticum, and Th. junceum provide an excellent
opportunity for the transfer of a range of genes responsible
*Correspondence: [email protected]
554
for resistance to pathogens and abiotic stress. However,
various levels of disturbance in the chromosome pairing
of newly formed hybrids may influence their genome
stability, seed set, and perenniality (Turner et al., 2013).
Since the initial hybridization between wheatgrasses
and wheat about 70 years ago (Armstrong, 1936), a
number of genes for resistance against wheat diseases have
been transferred into wheat in the form of wheat–alien
chromosome translocations. Thinopyrum intermedium
(Host) Barkworth and Dewey (2n = 6x = 42) [syn. Agropyron
intermedium (Host) Beauvoir; syn. Elytrigia intermedia
(Host) Nevski] has frequently been used in crosses aimed
at the transfer of genes for efficient resistance to rusts
(Lr38: Friebe et al., 1992; Sr44: Friebe et al., 1996; Turner et
al., 2013), powdery mildew (Pm40: Luo et al., 2009; Pm43:
He et al., 2009), barley yellow dwarf virus (Bdv2: Zhang
et al., 1992; Bdv3: Ohm et al., 2005), and wheat streak
mosaic virus (Wsm1: Friebe et al., 2009), as summarized
by Li and Wang (2009). Based on in situ hybridization
data, this allohexaploid species was found by Chen et al.
(1998) to consist of the genomes JJJsJsSS. Although this
formula varies in the work of different authors, there is
general agreement that it consists of 2 very similar (J = E
and St) genomes plus a third genome of unknown origin.
Chen’s genomic formula was contested by other authors,
GEORGIEVA et al. / Turk J Biol
including Mahelka et al. (2011), who analyzed chloroplast
trn L-F and partial nuclear GBSSI sequences, performed
multicolor GISH (mcGISH), and revealed new aspects of
the genomic composition of Th. intermedium. The authors
concluded that while Pseudoroegneria is the most probable
maternal parent of the accessions analyzed, nuclear GBSSI
sequences suggested the contribution of distinct lineages
corresponding to the following present-day genera:
Pseudoroegneria, Dasypyrum, Taeniatherum, Aegilops,
and Thinopyrum. These results showed that a high level of
heterogeneity existed at the genomic level in this species.
Sequential fluorescence and genomic in situ
hybridization (FISH and GISH) on Th. intermedium
chromosomes using the DNAs of microdissected
chromosomes indicated that the Jst genome is closer to the
J/E genomes from Th. bessarabicum and Th. elongatum
than to the introgressed St genome centromeric sequences
(Deng et al., 2013).
Several bread wheat–Th. intermedium amphiploids
have been obtained for leaf rust, yellow rust, and powdery
mildew resistance (Friebe et al., 1996; Bao et al., 2009; He
et al., 2009; Luo et al., 2009; Chang et al., 2010; Zeng et al.,
2013), as well as high protein content (Han et al., 2004;
Georgieva et al., 2011b). The aim of the present work was
to combine the FISH and GISH techniques in order to
characterize a new wheat–Th. intermedium introgression
line 29(1-57) with the ultimate goal of utilizing it for wheat
improvement.
2. Materials and methods
2.1. Plant material and chromosome preparation
Line 29(1-57) was obtained from a cross between T.
aestivum cv. Russalka as the female parent and Th.
intermedium (accession of unknown origin) as the male
parent, followed by backcrossing with the hexaploid
wheat cv. Sadovo. Line 29(1-57) is fertile and intermediate
in morphology between both original parents. Roottip chromosome spreads of the line were prepared as
described by Lukaszewski et al. (2004).
2.2. DNA probes and labeling
Several DNA probes were used for in situ hybridization: (1)
total genomic DNA of Th. intermedium; (2) total genomic
DNA of Th. bessarabicum (J genome) and Pseudoroegneria
spicata (St genome); (3) clone pTa71, containing the
18S-5.8S-26S rDNA repeat unit from T. aestivum (Gerlach
and Bedbrook, 1979); (4) Afa-family (Nagaki et al., 1995),
a subclone of the pAs1 tandem repetitive sequences
originating from Ae. squarrosa L. (Rayburn and Gill, 1986);
and (5) clone pSc119.2, containing the 120 bp repeat unit
of a tandemly arranged DNA family derived from Secale
cereale (McIntyre et al., 1990).
The probes were labeled with digoxigenin-11-dUTP or
biotin-11-dUTP using nick translation (pTa71), random
priming (genomic DNA), or the polymerase chain reaction
(Afa-family and pSc119.2) with universal forward and
reverse primers.
For GISH analysis, the genomic DNA probes of
Th. intermedium and J and St genome were added to
the hybridization mixture at ratios of 1:35 and 1:50,
respectively, with the nonlabeled blocking genomic wheat
DNA.
2.3. In situ hybridization
In situ hybridization was performed as described by Sepsi
et al. (2008). Digoxigenin-labeled probes were detected
using antidigoxigenin-rhodamine Fab fragments (Roche
Diagnostics, GmbH), while biotin-labeled probes were
detected using streptavidin-fluorescein thiocyanate
(FITC) (Roche Diagnostics, GmbH). All the preparations
were counterstained with 1 µg/mL 4’,6-diamidino2-phenylindole (DAPI) and mounted in Vectashield
antifade solution (Vector Laboratories, Ltd). The best cells
were captured with a Zeiss Axioscope 2 Epifluorescence
microscope equipped with a Spot CCD camera (Diagnostic
Instruments, Inc., USA) and Image Pro Plus 5.1 software
(Media Cybernetics, USA).
2.4. Determination of crude protein content (CPC)
The seed protein content was assessed according to the
Kjeldahl procedure described by Georgieva et al. (2011b).
The summarized crude protein data are reported as mean
values ± standard error (SE) for each group investigated
[Sadovo and 29(1-57)]. Differences between the 2
groups were assessed for statistical significance with the
parametric Student’s t-test. Data analysis was performed
with SPSS v. 15.0. A P-value less than 0.05 was considered
statistically significant.
3. Results
Thinopyrum intermedium is commonly used for hay
production and pasture. Crude protein measurement
plays a role in livestock feed, and it estimates the amount
of protein calculated from the determined nitrogen
content. Overall, the introgression line 29(1-57) showed a
tall wheatgrass-like phenotype (Figure 1). Follow-up data
evaluations of crude protein content were available for
29(1-57) (7-year data). Significant differences (P < 0.001)
in crude protein content were found between the control
wheat and the introgression line. The crude protein
content in the seeds was about 19.13 ± 2.4% for 29(1-57),
compared with 12.39 ± 2.09% for the standard cultivar,
Sadovo; in the introgression line the crude protein content
increased significantly.
The chromosome number of 29(1-57) varied from
40 to 42. The GISH technique was used to determine
the number of Th. intermedium chromosomes in the
introgression line. By probing with digoxigenin-labeled
total genomic DNA of Th. intermedium and blocking with
555
GEORGIEVA et al. / Turk J Biol
Figure 1. Spike and seed morphology of wheat–wheatgrass line
29(1-57).
wheat genomic DNA, 14 Th. intermedium chromosomes
and 28 wheat chromosomes were distinguished (Figure
2A). The mcGISH technique, using J and St genomic
DNA as probes, allowed us to examine the chromosome
composition of the alien species. Six chromosomes (Al2Al7) were labeled at the terminal regions with a stronger
red fluorescence signal and were weakly hybridized over
the remaining parts of the chromosomes (Figure 2B). J
and St genomic DNA also hybridized to the other parts,
resulting in yellowish hybridization signals; therefore, they
were more likely to belong to the J genome than St or Jst.
There were strong red hybridization signals in 2
chromosomes (Al1), and they were identified as St-genome
chromosomes (Figure 2B). There were no Jst chromosomes
(where the centromeric region was hybridizing with St
genomic DNA) present in 29(1-57). No translocation
events involving wheat and Th. intermedium chromosomes
were observed.
The use of sequential three-color FISH made it
possible to distinguish and characterize the wheat and Th.
intermedium chromosomes in 29(1-57). The simultaneous
hybridization of the repetitive DNA probes pSc119.2, Afafamily, and pTa71 allowed the wheat chromosome to be
identified unequivocally (Figure 3A). The chromosomes
3A, 4A, 1B, 3B, 4B, 5B, 6B, 7B, 1D, 2D, 3D, 5D, 6D, and 7D
were identified; the wheat chromosome pairs 1A, 2A, 5A,
6A, 7A, 2B, and 4D were not detected (Figure 3A).
The 7 alien chromosome pairs in 29(1-57) were
distinguished with sequential FISH/GISH (Figure 3) based
on the pattern and intensity of the Afa-family, pSc119.2,
Figure 2. GISH patterns of 29(1-57). (A) Total genomic DNA of Th. intermedium was used for probing. Fourteen wheatgrass
chromosomes fluoresced in red (marked Al as alien), wheat chromosomes were unlabeled. (B) Multicolor genomic in situ hybridization
with St (Pseudoroegneria spicata) and J (Thinopyrum bessarabicum) genomic DNA. Scale bar: 10 µm.
556
GEORGIEVA et al. / Turk J Biol
Figure 3. (A) FISH karyotype of metaphase chromosomes of amphiploid 29(1-57). The repetitive DNA probes
were labeled using biotin-11-dUTP and digoxigenin-11-dUTP and detected with streptavidin-FITC and
antidigoxigenin-rhodamine, respectively: pScll9.2 (green), Afa-family probe (red), pTa71 (yellow). Scale bar:
10 µm. (B) FISH pattern of the seven types of Th. intermedium chromosomes in the wheat–Th. intermedium
amphiploid 29(1-57). (C) Comprehensive FISH ideogram representation of alien introgressions.
and pTa71 hybridization sites, chromosome size, and arm
ratio. To facilitate identification, the alien chromosomes in
29(1-57) were designated as Al1-7. All the Al chromosome
pairs were metacentric to nearly submetacentric (Figure
3B). A karyogram was constructed based on the metaphase
images of the wheatgrass chromosomes (Figure 3B), and it
provided an overview of the specific FISH patterns of the
Th. intermedium chromosomes (Figure 3C). Probe pTa71
gave an atypical hybridization signal on alien chromosome
Al2, which was located near consecutive pSc119.2 and
Afa-family signals.
4. Discussion
Wild relatives of bread wheat have been extensively used
as valuable sources for the introgression of useful traits
for the purpose of wheat improvement. Thinopyrum
intermedium is a donor of a number of useful genes for
wheat improvement, such as resistance to pests, rust
diseases, and tolerance to abiotic stresses (e.g., salinity and
drought). Like its progenitor, the introgression line 29(157) displayed increased resistance to leaf rust, yellow rust,
and powdery mildew compared to the wheat cultivar and
other wheat–Thinopyrum stabilized hybrids under field
conditions (Tyankova, 2011).
Partial amphiploids between wild species and wheat
are often cytologically stable at 2n = 42 or 56 (BrasileiroVidal et al., 2005). This is due to normal meiotic pairing
and recombination between genomes in the F1 hybrids
or in their subsequent selfed or back-crossed progenies
(Cifuentes and Benavente, 2009). Partial amphiploids have
been reported to contain 42 (38–42) wheat and 14 (14–18)
alien chromosomes (Fedak and Han, 2005). In the present
study the introgression line possesses 42 chromosomes: 28
from wheat and 14 from Th. intermedium. This chromosome
number has been detected in other wheat lines with
introgressed Thinopyrum chromatin but usually refers to
557
GEORGIEVA et al. / Turk J Biol
substitution lines containing 1 alien pair, such as P12, P29,
and Zhong 199 (Sharma et al., 1999; Li et al., 2014).
Usually, efficient gene transfer between wheat
and Thinopyrum species in the form of chromosome
substitutions or translocation lines involves the D genome,
and only rarely the A or B genomes (Qi et al., 2007), because
the wheatgrass genome shares more homology with the D
genome (Liu et al., 2007). In the present study, the 29(157) introgression line differs from those obtained so far,
because it contains 14 alien chromosomes involving the
replacement of five chromosome pairs from the A genome
and 1 chromosome pair each from the B or D genomes
of the wheat parent. Compared with previous studies the
atypical FISH pattern observed for alien chromosome
Al2, especially the pattern of pTa71 probe (Li et al., 2004;
Georgieva et al., 2011a, 2011b), suggests that reciprocal
translocations may exist between the J and St genomes.
GISH has been used to characterize genomes and
chromosomes in hybrid polyploids, hybrid lines, partial
allopolyploids, and recombinant breeding lines and to
localize and detect alien introgressions (Chen et al., 2003;
Tek et al., 2004; Fedak and Han, 2005; Bao et al., 2009;
Chang et al., 2010; Georgieva et al., 2011b; Xie et al., 2013;
Książczyk et al., 2015). In the present study, GISH using
Thinopyrum intermedium, St, and J genomic DNA as
probes provided information about the number and type
of introgressed alien chromosomes and demonstrated the
presence of intergenomic chromosome rearrangements.
The results obtained with GISH were in agreement with
those of the FISH analysis and indicated that chromosomes
with different, unknown patterns were of alien origin. St
genomic DNA also hybridized to the NOR region of the
wheat chromosomes (Figure 2B), while Th. intermedium
DNA did not hybridize to the satellite regions of wheat
(Figure 2A), despite the fact that Th. intermedium also
contains St genomes. Tang et al. (2000) also found that it is
possible to distinguish the 3 satellited chromosomes of Th.
intermedium by GISH analysis using genomic DNA from
Ps. strigosa as a probe (Ps. spicata and Ps. strigosa also have
St genomes). In their study the probe hybridized to the
inner border of the satellite on the short arm.
Compared to the FISH karyotype of E genome
published by Linc et al. (2012), Al2 had a similar FISH
pattern to 6E, Al3 to 3E, and Al7 to 7E chromosomes of
different Elytrigia elongata accessions. Al1 was the shortest
alien chromosome. This was in good agreement with other
publications (Hsiao et al., 1986; Chen et al., 1999), as St
chromosomes were shorter than J chromosomes.
Chen et al. (1999) noted that the J genome
chromosomes present in TAF46 wheat × Th. intermedium
partial amphiploid displayed a unique GISH hybridization
pattern with the St genomic DNA probe, in which St
genomic DNA strongly hybridized at the terminal regions
and hybridized weakly over the remaining parts of the
chromosomes. The authors suggested this phenomenon
as a diagnostic marker for distinguishing J genome
chromosomes from Jst or St genome or wheat ABD genome
chromosomes.
In the present study sequential GISH and FISH were
used to discriminate the constituent genomes and to
identify the wheat and wheatgrass chromosomes in an
introgression line containing 42 chromosomes. Fourteen
wheat chromosomes (mainly A genome) were substituted
by 14 wheatgrass chromosomes. GISH revealed the
number of introgressed Th. intermedium chromosomes,
whereas FISH-banding allowed the chromosomes
belonging to both parents to be accurately identified. As
a genetic source for improving disease resistance and
quality, the 29(1-57) line could be a promising crossing
partner in wheat breeding programs. However, further
insight into the detailed genetic constitution of this line
will be required before initiation of a crossing program.
Acknowledgments
This
work
was
supported
by
grant
no.
BG051PO001-3.3.06-0025, financed by the European
Social Fund and Operational Program Human Resources
Development (2007–2013), and co-financed by the
Bulgarian Ministry of Education and Science and the
Hungarian National Scientific Research Fund (OTKA
K104382, OTKA K108555). Thanks are due to B Hooper
for revising the manuscript linguistically.
References
Armstrong JM (1936). Hybridization of Triticum and Agropyron,
crossing results and description of the first generation hybrids.
Can J Res 14: 190–202.
Bao Y, Li X, Liu S, Cui F, Wang H (2009). Molecular cytogenetic
characterization of a new wheat–Thinopyrum intermedium
partial amphiploid resistant to powdery mildew and stripe
rust. Cytogenet Genome Res 126: 390–395.
558
Brasileiro-Vidal AC, Cuadrado A, Brammer S, Benko-Iseppon AM,
Guerra M (2005). Molecular cytogenetic characterization
of parental genomes in the partial amphidiploid Triticum
aestivum × Thinopyrum ponticum. Genet Mol Biol 28: 308–313. Chang ZJ, Zhang XJ, Yang ZJ, Zhan HX, Li X, Liu C, Zhang CZ
(2010). Characterization of a partial wheat–Thinopyrum
intermedium amphiploid and its reaction to fungal diseases of
wheat. Hereditas 147: 304–312.
GEORGIEVA et al. / Turk J Biol
Chen P, Zhou B, Qi L, Liu D (1995). Identification of wheat-Haynaldia
villosa amphiploid, addition, substitution and translocation
lines by in situ hybridization using biotin-labelled genomic
DNA as a probe. Acta Genet Sinica 22: 380–386.
Chen Q, Conner RL, Laroche A, Ji WQ, Armstrong KC, Fedak G
(1999). Genomic in situ hybridization analysis of Thinopyrum
chromatin in a wheat – Th. intermedium partial amphiploid
and six derived chromosome addition lines. Genome 42:
1217–1223.
Chen Q, Conner RL, Laroche A, Thomas JB (1998). Genome analysis
of Thinopyrum intermedium and Thinopyrum ponticum using
genomic in situ hybridization. Genome 41: 580–586.
Chen Q, Conner RL, Sun SC, Ahmad F, Laroche A, Graf RJ (2003).
Molecular cytogenetic discrimination and reaction to Wheat
streak mosaic virus and the wheat curl mite in Zhong series of
wheat–Thinopyrum intermedium partial amphiploids. Genome
46: 135–145.
Cifuentes M, Benavente E (2009). Wheat–alien metaphase I pairing
of individual wheat genomes and D genome chromosomes in
interspecific hybrids between Triticum aestivum L. and Aegilops
geniculata Roth. Theor Appl Genet 119: 805–813.
Deng C, Bai L, Fu S, Yin W, Zhang Y, Chen Y, Wang R, Zhang X, Han
F, Hu Z (2013). Microdissection and chromosome painting of
the alien chromosome in an addition line of wheat-Thinopyrum
intermedium. PLoS ONE 8: e72564.
Farkas A, Molnár I, Dulai S, Rapi S, Oldal V, Cseh A, Kruppa K,
Molnár-Láng M (2014). Increased micronutrient content (Zn,
Mn) in the 3Mb(4B) wheat–Aegilops biuncialis substitution and
3Mb.4BS translocation identified by GISH and FISH. Genome
57: 61–67.
Fedak G, Han F (2005). Characterization of derivatives from wheat–
Thinopyrum wide crosses. Cytogenet Genome Res 109: 360–
367.
Friebe B, Jiang J, Raupp WJ, McIntosh RA, Gill BS (1996).
Characterization of wheat-alien translocations conferring
resistance to diseases and pests: current status. Euphytica 91:
59–87.
Gerlach WL, Bedbrook JR (1979). Cloning and characterization of
ribosomal RNA genes from wheat and barley. Nucleic Acids
Res 7: 1869–1885.
Han F, Liu B, Fedak G, Liu Z (2004). Genomic constitution and
variation in five partial amphiploids of wheat–Thinopyrum
intermedium as revealed by GISH, multicolor GISH and seed
storage protein analysis. Theor Appl Genet 109: 1070–1076.
He RL, Chang ZJ, Yang ZJ, Yuan ZY, Zhan HX, Zhang XJ, Liu JX
(2009). Inheritance and mapping of powdery mildew resistance
gene Pm43 introgressed from Thinopyrum intermedium into
wheat. Theor Appl Genet 118: 1173–1180.
Hsiao C, Wang RRC, Dewey DR (1986). Karyotype analysis and
genome relationships of 22 diploid species in the tribe Triticeae.
Can J Genet Cytol 28: 109–120.
Kumar S, Friebe B, Gill BS (2010). Fate of Aegilops speltoides-derived,
repetitive DNA sequences in diploid Aegilops species, wheatAegilops amphiploids and derived chromosome addition lines.
Cytogenet Genome Res 129: 47–54.
Książczyk T, Zwierzykowska E, Molik K, Taciak M, Krajewski P,
Zwierzykowski Z (2015). Genome-dependent chromosome
dynamics in three successive generations of the allotetraploid
Festuca pratensis × Lolium perenne hybrid. Protoplasma 252:
985–896.
Li DY, Ru YY, Zhang XY (2004). Chromosomal distribution of the
18S-5.8S-26S rDNA loci and heterogeneity of nuclear ITS
regions in Thinopyrum intermedium (Poaceae: Triticeae). Acta
Bot Sin 46: 1234–1241.
Li H, Wang X (2009). Thinopyrum ponticum and Th. intermedium:
the promising source of resistance to fungal and viral diseases
of wheat. J Genet Genomics 36: 557–565.
Li XJ, Li G, Jiang XL, Sun Y, Feng SW, Hu TZ, Ru ZG, Zhang
LL, Song J (2014). Identification of a wheat–Thinopyrum
intermedium substitution line as revealed by GISH, FISH, SSR
and EST-SSR markers. Research on Crops 15: 345–351.
Linc G, Sepsi A, Molnár-Láng M (2012). A FISH karyotype to study
chromosome polymorphisms for the Elytrigia elongata E
genome. Cytogenet Genome Res 136: 138–144.
Friebe B, Qi LL, Wilson DL, Chang ZJ, Seifers DL, Martin TJ, Fritz
AK, Gill BS (2009). Wheat-Thinopyrum intermedium
recombinants resistant to wheat streak mosaic virus and
Triticum mosaic virus. Crop Sci 49: 1221–1226.
Liu W, Seifers DL, Qi LL, Friebe B, Gill BS (2011). A compensating
wheat-Thinopyrum intermedium Robertsonian translocation
conferring resistance to wheat streak mosaic virus and Triticum
mosaic virus. Crop Sci 51: 2382–2390.
Friebe B, Zeller FJ, Mukai Y, Forster BP, Bartos P, McIntosh RA
(1992). Characterization of rust-resistant wheat-Agropyron
intermedium derivatives by C-banding, in situ hybridization
and isozyme analysis. Theor Appl Genet 83: 775–782.
Liu Z, Li D, Zhang X (2007). Genetic relationships among five basic
genomes St, E, A, B and D in Triticeae revealed by genomic
Southern and in situ hybridization. J Integr Plant Biol 49:
1080–1086.
Georgieva M, Sepsi A, Molnár-Láng M, Tyankova N (2011a).
Molecular cytogenetic analysis of Triticum aestivum and
Thinopyrum intermedium using the FISH technique. C R Acad
Bulg Sci 64: 1713–1718.
Lukaszewski AJ, Rybka K, Korzun V, Malyshev SV, Lapinski
B, Whitkus R (2004). Genetic and physical mapping of
homoeologous recombination points involving wheat
chromosome 2B and rye chromosome 2R. Genome 47: 36–45.
Georgieva M, Sepsi A, Tyankova N, Molnár-Láng M (2011b).
Molecular cytogenetic characterization of two high protein
wheat–Thinopyrum intermedium partial amphiploids. J Appl
Genet 52: 269–277.
Luo PG, Luo HY, Chang ZJ, Zhang HY, Zhang M, Ren ZL (2009).
Characterization and chromosomal location of Pm40 in
common wheat: a new gene for resistance to powdery mildew
derived from Elytrigia intermedium. Theor Appl Genet 118:
1059–1064.
559
GEORGIEVA et al. / Turk J Biol
Mahelka V, Kopecky D, Pastova L (2011). On the genome constitution
and evolution of intermediate wheatgrass (Thinopyrum
intermedium: Poaceae, Triticeae). BMC Evol Biol 11: 127.
McIntyre CL, Pereira S, Moran LB, Appels R (1990). New Secale
cereale (rye) DNA derivatives for the detection of rye
chromosome segments in wheat. Genome 33: 635–640.
Molnár-Láng M, Linc G, Szakács É (2014). Wheat-barley
hybridization: the last 40 years. Euphytica 195: 315–329.
Munns R, James RA, Islam A, Colmer TD (2011). Hordeum
marinum-wheat amphiploids maintain higher leaf K+:Na+ and
suffer less leaf injury than wheat parents in saline conditions.
Plant and Soil 348: 365–377.
Nagaki K, Tsujimoto H, Isono K, Sasakuma T (1995). Molecular
characterization of a tandem repeat, Afa family, and
distribution among Triticeae. Genome 38: 479–486.
Ohm HW, Anderson JM, Sharma HC, Ayala L, Thompson N, Uphaus
JJ (2005). Registration of Yellow dwarf virus resistant wheat
germplasm line P961341. Crop Sci 45: 805–806.
Priya AM, Krishnan SR, Ramesh M (2015). Ploidy stability of Oryza
sativa. L cv IR64 transformed with the moth bean P5CS gene
with significant tolerance against drought and salinity. Turk J
Biol 39: 407–416.
Qi L, Friebe B, Zhang P, Gill BS (2007). Homoeologous
recombination, chromosome engineering and crop
improvement. Chromosome Res 15: 3–19.
Rayburn AL, Gill BS (1986). Molecular identification of the
D-genome chromosomes of wheat. J Hered 77: 253–255.
Schneider A, Linc G, Molnár I, Molnár-Láng M (2005). Molecular
cytogenetic characterization of Aegilops biuncialis and its use
for the identification of 5 derived wheat-Aegilops biuncialis
disomic addition lines. Genome 48: 1070–1082.
Sepsi A, Molnár I, Szalay D, Molnár-Láng M (2008). Characterization
of a leaf rust-resistant wheat-Thinopyrum ponticum partial
amphiploid BE-1, using sequential multicolor GISH and FISH.
Theor Appl Genet 116: 825–834.
Sharma H, Francki M, Crasta O, Gyulai G, Bucholtz D, Ohm H,
Anderson J, Perry K, Patterso F (1999). Cytological and
molecular characterization of wheat lines with Thinopyrum
intermedium chromosome additions, substitutions and
translocations resistant to Barley yellow dwarf virus. Cytologia
64: 93–100.
560
Tang S, Li Z, Jia X, Larkin PJ (2000). Genomic in situ hybridization
(GISH) analyses of Thinopyrum intermedium, its partial
amphiploid Zhong 5, and disease-resistant derivatives in
wheat. Theor Appl Genet 100: 344–352.
Tang ZX, Fu SL, Ren ZL, Zhou JP, Yan BJ, Zhang HQ (2008).
Variations of tandem repeat, regulatory element, and promoter
regions revealed by wheat-rye amphiploids. Genome 51: 399–
408.
Tek AL, Stevenson WR, Helgeson JP, Jiang J (2004). Transfer of tuber
soft rot and early blight resistances from Solanum brevidens
into cultivated potato. Theor Appl Genet 109: 249–254.
Turner MK, DeHaan LR, Jin Y, Anderson JA (2013). Wheatgrass–
wheat partial amphiploids as a novel source of stem rust and
Fusarium head blight resistance. Crop Sci 53: 1194–2005.
Tyankova ND (2011). Development and characteristics of wheatThinopyrum intermedium amphiploids. C R Acad Bulg Sci 64:
361–368.
Wang Y, Yu K, Xie Q, Kang H, Lin L, Fan X, Sha L, Zhang H, Zhou
Y (2011). Cytogenetic, genomic in situ hybridization (GISH)
and agronomic characterization of alien addition lines derived
from wheat-Psathyrostachys huashanica. Afr J Biotechnol 10:
2201–2211.
Whitford R, Fleury D, Reif JC, Garcia M, Okada T, Korzun V,
Langridge P (2013). Hybrid breeding in wheat: technologies to
improve hybrid wheat seed production. J Exp Bot 64: 5411–
5428.
Xie S, Ramanna MS, Visser RGF, Arens P, van Tuyl JM (2013).
Elucidation of intergenomic recombination and chromosome
translocation: meiotic evidence from interspecific hybrids of
Lilium through GISH analysis. Euphytica 194: 361–370.
Yang ZJ, Li GR, Feng J, Jiang HR, Ren ZL (2005). Molecular
cytogenetic characterization and disease resistance observation
of wheat-Dasypyrum breviaristatum partial amphiploid and its
derivatives. Hereditas 142: 80–85.
Zeng J, Cao W, Fedak G, Sun S, McCallum B, Fetch T, Xue A, Zhou
Y (2013). Molecular cytological characterization of two novel
durum-Thinopyrum intermedium partial amphiploids with
resistance to leaf rust, stem rust and Fusarium head blight.
Hereditas 150: 10–16.