Journal of Heredity 2012:103(4):594–597
doi:10.1093/jhered/ess009
Advance Access publication May 8, 2012
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Genetic Control of White Flower Color
in Scarlet Rosemallow (Hibiscus
coccineus Walter)
LYN A. GETTYS
From the University of Florida Institute of Food and Agricultural Sciences, Center for Aquatic and Invasive Plants, 7922
Northwest 71 Street, Gainesville, FL 32653. Gettys is now at the Department of Agronomy, University of Florida Institute
of Food and Agricultural Sciences, Fort Lauderdale Research and Education Center, 3205 College Avenue, Davie, FL 33314.
Address correspondence to Lyn A. Gettys at the address above, or e-mail: [email protected].
Scarlet rosemallow (Hibiscus coccineus Walter) is a diploid,
perennial, erect, and woody shrub. The species is a desirable
inclusion in home landscapes because it is a native plant with
attractive flowers and unusual foliage. The objective of
these experiments was to determine the number of loci,
number of alleles, and gene action controlling flower color
(red vs. white) in scarlet rosemallow. Three white-flowered
and 1 red-flowered parental lines were used to create S1
and F1 populations, which were self-pollinated or backcrossed to generate S2, F2, and BC1 populations. Evaluation
of these generations showed that flower color in these
populations was controlled by a single diallelic locus with
red flower color completely dominant to white. I propose
that this locus be named ‘‘white flower’’ with alleles
W and w.
Key words: complete dominance, hibiscus, native plant, single
diallelic locus
Scarlet rosemallow (Hibiscus coccineus Walter) is diploid (2n 5
2x 5 38) (Wise and Menzel 1971) and is one of three
species within section Muenchhusia (Heister ex Fabricium) of
the North American genus Hibiscus L., which comprises 5
species (Small 2004). Synonyms include H. coccineus var.
integrifolius, H. integrifolius, H. semilobatus, H. carolinianus, and
H. speciosus (Wunderlin and Hansen 2008), and other
common names include Texas star, wild red mallow, swamp
hibiscus, and swamp mallow. Scarlet rosemallow is
a perennial, deciduous, erect, and woody shrub that inhabits
swamps, marshes, sloughs, lake banks, and ditches from
southeastern Georgia, Alabama, and Texas to southern
Florida (Godfrey and Wooten 1979). Growth of the species
is typically vase-like with multiple erect stems that can grow
to 3 m in height. Juvenile leaves are deltoid and mostly
unlobed, whereas mature leaves are strongly palmate, with
deep clefts separating each leaf into 5 lobes (Godfrey and
Wooten 1979). Scarlet rosemallow is classified as an obligate
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wetland plant (USDA NRCS 2011), but performs well in
less inundated areas provided soil moisture is maintained at
adequate levels. Plants begin to flower in May in northern
Florida and bear brilliant red 5-petaled flowers that may be
as broad as 20 cm. Flowering continues until plants die back
in November or December. Plants remain dormant throughout the winter and begin to resprout from underground
rhizomes as early as March. Some sources list the species as
being hardy in United States Department of Agriculture
(USDA) Zones 7–11, whereas others suggest that the plant
can survive in regions as cold as Zone 5 (Odenwald and
Turner 2006; Gilman 2007; Anonymous 2011).
The perennial nature, native status, wide geographic
range, and showy flowers of scarlet rosemallow have made
the species a popular garden plant that is sometimes
available locally from selected native plant nurseries. In the
absence of local outlets, plant and seeds can easily be
purchased from any number of nurseries and private
individuals on the Internet. Although feral populations of
scarlet rosemallow almost exclusively bear red flowers,
white-flowered specimens are available in the nursery trade
and on the Internet. Variations in flower color are frequently
transmitted and inherited in a simple Mendelian fashion
(Durbin et al. 2003). For example, flower color variation is
controlled by a single diallelic locus in pickerelweed (Gettys
and Wofford 2007), stokes aster (Gaus et al. 2003), morning
glory (Zufall and Rauscher 2003), Chinese houses (Lankinen
2009), and safflower (Hamdan et al. 2008). However, the
genetic control of flower color in other species is more
complex. For example, Yue et al. (2008) found that lemon
ray flower color in sunflower was due to a recessive
condition at both of two independent loci, whereas
Griesbach (1996) discovered that 4 loci contributed to
flower color in petunia.
There is no published information outlining the genetic
control and inheritance of flower color in scarlet rosemallow. The objective of these experiments was to identify
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Brief Communications
Journal of Heredity
the type of gene action and number of loci controlling
flower color in these populations of scarlet rosemallow.
Materials and Methods
Plants used in these experiments were part of a population
maintained for breeding and genetic studies at the University
of Florida Center for Aquatic and Invasive Plants in
Gainesville. Parent, S1, and F1 populations were grown in 4-l
nursery containers, whereas F2, S2, and BC1 populations
were grown in 1-l nursery containers. All containers were
filled with a commercial potting mix amended with 10 or 5 g
(for 4-l and 1-l containers, respectively) of controlled-release
fertilizer per container. Plants were maintained in pollinatorfree greenhouses under ambient temperature conditions,
with supplemental lighting used to artificially increase day
length to 16 h. Young plants were irrigated using an
overhead mist system and were later transferred to
a subirrigation regime to ensure that plants received
adequate water. Plants were supported with bamboo stakes
as needed throughout the growing period.
Three white-flowered parents (coded W1, W2, and W3)
and 1 red-flowered plant (coded R) were utilized in these
experiments. All white-flowered parents were started from
seeds purchased from a hobbyist gardener in Texas via the
Internet and are presumably derived from the same
population. The red-flowered parent was a seedling from
a plant originally collected from a native population of
scarlet rosemallows at Lake George in central Florida and
maintained as part of an aquatic ornamentals collection in
Gainesville. Given the geographic separation of the sources
of the white-flowered parents and the red-flowered parent, it
is unlikely that the color morphs used as parents in these
experiments share close common ancestors. Each parent
was self-pollinated to create the S1 families W1 5, W2 5,
W3 5, and R 5. Cross- and reciprocal pollinations
were performed between parents to create the F1 families
W1 R, W2 R, and W3 R. Subsamples were selected
from each S1 and F1 family and were self-pollinated to
produce S2 and F2 families, respectively. In addition,
subsamples of F1 families were backcrossed to white-flowered
parents to generate BC1 families. Self, cross-, and reciprocal
pollinations to create S1 and F1 populations were performed
between June and September 2008, whereas self-pollinations
of S1 and F1 plants (to create S2 and F2 populations,
respectively) and backcrosses (to create BC1 populations)
took place between August and November 2009.
Flowers of scarlet rosemallow remain open and stigmas
are receptive for a single day, so pollinations were
performed as soon as flowers were fully expanded (usually
between 7:30 and 9:30 am). Self-pollination of parents to
create S1 families revealed that plants were highly self-fertile,
so seed parents in F1 crosses were emasculated prior to
pollen transfer to avoid self-pollination. Parental information and date of pollination were recorded on a jewelry tag,
which was then hung gently on the pedicel of the pollinated
flower. Capsule formation was monitored throughout the
2
maturation process and ripe capsules were collected 30–40
days after pollination. Seeds were manually removed from
each capsule and placed in a labeled manila coin envelope
until seed set was completed in all pollinations. All seeds in
each envelope were sown on the surface of a common
container filled with a commercial potting mix and placed
under mist irrigation (irrigation events every 2 h from 8 AM
to 8 PM; duration of each event 3 min) in a greenhouse with
temperature maintained between 25 and 30 C. Germinated
seeds were transplanted into individual labeled cells in 612
cell flats and maintained under mist until seedlings were
approximately 15 cm tall. Seedlings were then transplanted
into 4-l (S1 and F1 populations) or 1-l containers (S2, F2, and
BC1 populations), subirrigated and moved to the greenhouses described above.
All plants were grown to reproductive maturity and
scored for flower color. No maternal effects were noted, so
data for each family were pooled within each cross/
reciprocal set. Data from F1 and S1 families were used to
develop a proposed model to explain the type of gene action
and number of loci controlling flower color in scarlet
rosemallow. This model was used to assign likely genotypes
to parents used in these experiments and the model was
then verified by analyses of F2 and BC1 populations. All data
were analyzed using goodness-of-fit (chi-square or v2) tests
(Steel et al. 1997). A test for heterogeneity of data collected
from F2 families from different crosses was employed to
determine whether it was appropriate to pool data for all F2
progeny; the same test was applied to data from BC1
families.
Results and Discussion
All S1 and S2 progeny in the families W1 5, W2 5, and W3
5 bore white flowers, whereas all S1 and S2 progeny from
the family R 5 bore red flowers. In addition, all progeny in
the 3 F1 families bore red flowers. The simplest model that
explains the progeny types recovered in these S1 and F1
families is one with a single diallelic locus, with red flower
color completely dominant to white. Genotypes were
assigned to all 4 parents using the proposed model and
segregation of S1 and F1 progeny; all white-flowered parents
were homozygous recessive (ww) and the red-flowered
parent was homozygous dominant (WW ).
Sixteen F1 plants (3, 5, and 8 F1 plants from the families
W1 R, W2 R, and W3 R, respectively) were selfpollinated to produce 16 F2 families. Each F2 family
produced progeny that segregated in a manner that was not
significantly different from a 3 red:1 white ratio (data not
shown). Heterogeneity chi-square analysis showed that all F2
progeny within each cross/reciprocal set were from
populations with identical genotypic constitutions (data
not shown); therefore, data for F2 progeny within each
cross/reciprocal set were pooled. These pooled progeny
segregated in a manner that was not significantly different
from the expected 3 red:1 white ratio (Table 1). Heterogeneity chi-square analysis was performed on progeny from all
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Journal of Heredity 2012: 103(4)
Brief Communication
Table 1 Classification of plants of scarlet rosemallow in the F1, F2, and BC generations for red or white flower color
Parents
W1
W1
W1
W2
W2
W2
W3
W3
W3
R
R
R
R
R
R
R
R
R
Generation Genotype
F1
F2
BC1
F1
F2
BC1
F1
F2
BC1
Number of plants observed
Expected ratio
Number of plants expected
Red
White
Red
White
Red
White
v2
P
0
24
36
0
56
35
0
73
168
—
3
1
—
3
1
—
3
1
—
1
1
—
1
1
—
1
1
—
84.00
40.0
—
177.00
38.0
—
194.25
161.5
—
28.00
40.0
—
59.00
38.0
—
64.75
161.5
—
0.76
0.80
—
0.20
0.47
—
1.40
0.79
—
0.38
0.37
—
0.65
0.49
—
0.24
0.37
ww WW 11
Ww 5
88
ww Ww
44
ww WW 40
Ww 5
180
ww Ww
41
ww WW 49
Ww 5
186
ww Ww 155
Cross- and reciprocal pollinations are pooled within each parental set and are listed by the cross (e.g., observations attributed to W1 R include data from
W1 R and from R W1).
F2 families and the test was not significant; therefore, data
for all segregating F2 progeny from all F1 families were
pooled and subjected to chi-square analysis. These pooled
progeny segregated in a manner that was not significantly
different from the expected 3 red:1 white ratio (P 5 0.31).
Thirteen F1 plants (2, 2, and 9 F1 plants from the families
W1 R, W2 R, and W3 R, respectively) were
backcrossed to white-flowered parents to produce 13 BC1
families. Each BC1 family produced progeny that segregated in
a manner that was not significantly different from a 1 red:1
white ratio (data not shown). Heterogeneity chi-square analysis
showed that all BC1 progeny within each backcross/reciprocal
set were from populations with identical genotypic constitutions (data not shown); therefore, data for BC1 progeny within
each backcross/reciprocal set were pooled. These pooled
progeny segregated in a manner that was not significantly
different from the expected 1 red:1 white ratio (Table 1).
Heterogeneity chi-square analysis was performed on progeny
from all BC1 families and the test was not significant;
therefore, data for all segregating BC1 progeny were pooled
and subjected to chi-square analysis. These pooled progeny
segregated in a manner that was not significantly different
from the expected 1 red:1 white ratio (P 5 0.36).
Conclusion
The results of these experiments suggest that flower color in
these populations of scarlet rosemallow is controlled by 2
alleles at a single locus. Gene action is completely dominant,
with white flower color recessive to red. All progeny in these
experiments segregated as expected when tested against this
model. Self-pollination of white-flowered plants (genotype
ww) produced exclusively white-flowered progeny, whereas
self-pollination of red-flowered plants resulted in progeny
that only bore red flowers (parent genotype WW; source of
S1 and S2 populations) or progeny with red and white
flowers in a 3:1 ratio (parent genotype Ww; source of F2
populations). I propose that this locus controlling flower
color in scarlet rosemallow be named ‘‘white flower’’ with
alleles W and w.
The mechanism responsible for this novel flower color is
unknown and beyond the scope of this paper. However, it
seems likely that white flowers in scarlet rosemallow are the
result of a loss-of-function mutation affecting the anthocyanin pathway such as that described by Rauscher (2008).
Red flower color in wild-type scarlet rosemallow results
from a combination of the flavonoid aglycones quercetin
and cyanidin in an approximate 1:2 ratio (Puckhaber et al.
2002). Quercetin and cyanidin represent alternate pathways
that branch from DH-quercitin, an intermediate substrate
that is converted to one of these two aglycones (Figure 1).
DH-quercitin is converted to quercetin by the enzyme
flavonol synthase, whereas the substrate becomes cyanidin
in a 2-step process mediated by the enzymes dihydroflavonol 4-reductase and anthocyanidin synthase (Rauscher
2008). Although I did not investigate the biochemical
composition of floral pigments in this study, Puckhaber
et al. (2002) found that levels of quercetin were highest in
white-flowered species of Hibiscus.
Precursor
substrate
Quercetin
flavonol synthase
DH-quercitin
dihydroflavonol 4-reductase
anthocyanidin synthase
Cyanidin
Figure 1. Simplified diagram of the portion of the anthocyanin pathway that is likely responsible for flower color in scarlet
rosemallow. The precursor DH-quercitin is converted to either quercetin or cyanidin via opposing pathways.
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Journal of Heredity
Most transitions from pigmented to white flowers are
the result of loss-of-function mutations, which can arise in
a variety of ways. For example, mutations in regulatory
regions may damage or render inoperable transcription
factors and other elements that regulate and control gene
function. Coding regions may be disrupted through
insertions, deletions, or point mutations, which cause
frameshift mutations and alter the reading frame. The end
result of mutations that affect the components of the
anthocyanin pathway is loss of function in a branch of the
pathway. In scarlet rosemallow, it seems likely that the branch
responsible for conversion of the intermediate substrate
DH-quercitin to cyanidin is inoperable due to a mutation in
the gene coding for the enzyme dihydroflavonol 4-reductase.
This renders the branch leading to cyanidin synthesis
nonfunctional and excess DH-quercitin is diverted to the
alternate branch that produces quercetin instead.
Scarlet rosemallow is a desirable native perennial plant
that has great utility in wetland areas and in home gardens
and landscapes. The species has a number of attractive
characteristics that appeal to those searching for an attractive,
unusual, and woody perennial plant to include in vegetation
plans. Although red-flowered specimens of scarlet rosemallow can be locally common, white-flowered plants are not
always accessible to growers who wish to offer this color
variant of the species. This research revealed that white
flowers are the result of a recessive condition at the white
flower locus, so white-flowered plants of scarlet rosemallow
are inherently true breeding for flower color. Nurseries and
growers of native, ornamental, and wetland plants can exploit
this information to produce white-flowered plants for their
customers simply by maintaining white-flowered plants that
can be used as parents in seed production.
Funding
This research was supported by the University of Florida
Center for Aquatic and Invasive Plants and the Florida
Agricultural Experiment Station (FAES). Mention of a trademark or a proprietary product does not constitute a guarantee
or warranty of the product by the FAES nor does it imply its
approval to the exclusion of other products that may be
suitable.
Acknowledgments
I wish to thank William Haller, Margaret Glenn, Jody Strobel, Kyle Thayer,
Jeremy Douglas, and Kacie Ritter for their invaluable assistance with this
project. I also wish to dedicate this paper to the memory of my dear friend,
teacher, and mentor, the late Paul Pfahler.
References
Anonymous. 2011. Tall plants for gardens. [cited 2011 Aug 10]. Available
from: http://www.gardenguides.com/119479-tall-plants-gardens.html
Durbin ML, Lundy KE, Morrell PL, Torres-Martinez CL, Clegg MT. 2003.
Genes that determine flower color: the role of regulatory changes in the
evolution of phenotypic adaptations. Mol Phylogenet Evol. 29:507–517.
4
Gaus J, Werner D, Gettys L, Griesbach R. 2003. Genetics and biochemistry
of flower color in stokes aster. Acta Hortic. 624:449–453.
Gettys LA, Wofford DS. 2007. Inheritance of flower color in pickerelweed
(Pontederia cordata L.). J Hered. 98:629–632.
Gilman EF. 2007. Hibiscus coccineus swamp mallow. IFAS Publication No.
FPS-253. Gainesville (FL): University of Florida Institute of Food and
Agricultural Sciences; [cited 2011 Aug 10]. Available from: http://
edis.ifas.ufl.edu/fp253
Godfrey RK, Wooten JW. 1979. Aquatic and wetland plants of the
southeastern United States: dicotyledons. Athens (GA): The University of
Georgia Press.
Griesbach RJ. 1996. The inheritance of flower color in Petunia hybrida Vilm.
J Hered. 87:241–245.
Hamdan YAS, Begoña Pérez-Vich JM, Fernández-Martı́nez LV. 2008.
Inheritance of very high oleic acid content and its relationship with several
morphological and physiological traits. In: Knights SE, Potter TD, editors.
Safflower: unexploited potential and world adaptability. Proceedings of the
7th International Safflower Conference; Wagga Wagga, New South Wales
(Australia): Australian Oilseeds Federation; [cited 2011 Aug 10]. Available
from: http://www.australianoilseeds.com/__data/assets/pdf_file/0018/
6813/final_Velasco_oral_paper.pdf
Lankinen Å. 2009. Upper petal lip colour polymorphism in Collinisia
heterophylla (Plantaginaceae): genetic basis within a population and its use as
a genetic marker. J Genet. 88:205–215.
Odenwald NG, Turner JR. 2006. Identification, selection, and use of
southern plants: for landscape design. 4th ed. Baton Rouge (LA): Claitor’s
Law Books and Publishing.
Puckhaber LS, Stipanovic RD, Bost GA. 2002. Analyses for flavonoid
aglycones in fresh and preserved Hibiscus flowers. In: Janick J, Whipkey A,
editors. Trends in new crops and new uses. Proceedings of the 5th National
Symposium. New crops and new uses: strength in diversity. Alexandria
(VA): ASHS Press. p. 556–563 Available from: http://www.hort.purdue.
edu/newcrop/ncnu02/pdf/bost-556.pdf
Rauscher MD. 2008. Evolutionary transitions in floral color. Int J Plant Sci.
169:7–21.
Small RL. 2004. Phylogeny of Hibiscus sect. Muenchhusia (Malvaceae) based
on chloroplast rpL16 and ndhF, and nuclear ITS and GBSSI sequences. Syst
Bot. 29:385–392.
Steel RGD, Torrie JH, Dickey DA. 1997. Principles and procedures of
statistics: a biometrical approach. 3rd ed. New York: WCB McGraw-Hill.
USDA NRCS. 2011. The PLANTS database. Greensboro (NC): National
Plant Data Team; [cited 2011 Aug 10]. Available from: http://plants.usda.
gov/java/profile?symbol5HICO2
Wise DA, Menzel MY. 1971. Genetic affinities of the North American
species of Hibiscus sect. Trionum. Brittonia. 23:425–437.
Wunderlin RP, Hansen BF. 2008. Atlas of Florida vascular plants. Tampa
(FL): Institute for Systematic Botany, University of South Florida; [cited
2011 Aug 10]. Available from: http://www.florida.plantatlas.usf.edu/
Plant.aspx?id53189
Yue B, Vick BA, Yuan W, Hu J. 2008. Mapping one of the 2 genes
controlling lemon ray flower color in sunflower (Helianthus annuus L.).
J Hered. 99:564–567.
Zufall RA, Rauscher MD. 2003. The genetic basis of a flower color
polymorphism in the common morning glory (Ipomea purpurea). J Hered.
94:442–448.
Received August 11, 2011; Revised January 3, 2012;
Accepted January 22, 2012
Corresponding Editor: John Stommel
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