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Molecular Plant Breeding 2010, Vol.1 No.3
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Since 2006, we began to transfer the white grain color
gene
R-D1a
of CW565 into Chuanmai 42
by selecting
the heterozygous genotype on
R-D1
allele with SSR
markers Xgwm3 and Xgwm314 in the backcross
progeny, and several advanced white grained lines
with good agronomic traits as Chuanmai 42 were
developed. The results indicated that the two SSR
markers can be used in marker-assisted selection for
breeding white grain wheat variety by discarding the
red grain color gene or selecting the white grain color
gene in the progeny of SHWs or Chuanmai 42 crossed
with white grain color wheat.
3 Materials and methods
3.1 Plant materials
Two wheat lines, Chuanmai 42 with red grain color
and ChuanW565 (CW565) (
R-A1a, R-B1a
and
R-D1a
)
with white grain color were employed in the present
experiment. Both Chuanmai 42 and CW565 were
developed and provided by Crop Research Institute,
Sichuan Academy of Agricultural Sciences in Sichuan
province from China.
Chuanmai42 was developed using a red grain color
synthetic hexaploid wheat (Syn769) crossed and
backcrossed with two white grain color local wheat
varieties Sw3243 (
R-A1a, R-B1a
and
R-D1a
) and
Chuan6415 (
R-A1a, R-B1a
and
R-D1a
), respectively.
The synthetic hexaploid wheat Syn769 was developed
from a white grain color durum wheat Decoy1
(2n=4x=28, AABB) (
R-A1a
and
R-B1a
) and the red
grain color
Ae. tauschii
accession SQ-188 (2n=2x=14,
DD)(
R-D1b
) by CIMMYT. According to the pedigree,
the character of red grain color of Chuanmai42 should
be derived from the D genome donor SQ-188 of
Syn769, and the red grain color alleles conferring
Chuanmai42 are
R-A1a, R-B1a
and
R-D1b
.
The Chuanmai 42 was hybridized with CW565 under
field conditions in Chengdu city of Sichuan province
in 2004. For seed multiplication during the summer
season, the F
1
seeds were planted at Yunnan
Agricultural University in Kunming city. During the
winter season, the F
2
populations, obtained from
Yunnan Agricultural University, together with their
parents and F
1
’s were grown as individual plants for
analysis in the field of Chengdu city. A total of 1 015
F
2
plants were harvested individually for estimating
the grain color and analyzing the inheritance of the
character of red color of Chuanmai 42 in the spring of
2005. For confirming the recessive plants, the 241
recessive lines of F
2
population with white grain color
were selected and planted (30~40 plants each) during
the winter season. In the spring of 2006, the F
2-3
single
plants derived from 241 recessive lines were harvested
and used to estimate the grain color again.
The 1 015 plants of F
2
population of Chuanmai
42×CW565 was used for the inheritance of the red
grain trait, and 241 F
2
recessive lines confirmed by
it’s F
2-3
populations were used for the mapping of red
grain color gene.
3.2 Trait evaluation
At the completely mature time, pericarp color of single
plant F
2
populations and F
2-3
populations derived from
the recessive lines of Chuanmai42×CW565, which was
determined in grain color after single plant threshing.
The grain color was evaluated by using NaOH method
and visual assessments. For the NaOH method, thirty
to forty seeds of each line were placed into 100×15 mm
petri-dishes. A 5% NaOH solution was poured over
the seeds and they were soaked in solution overnight.
The NaOH solution gave red wheat a dark red color,
while white wheat assumes a straw yellow color.
3.3 DNA extraction and microsatellite analysis
Total genomic DNA of the parents and F
2
single
plants was extracted from leaf material of individual
plants using the CTAB protocol (Sharp et al., 1988).
According to results of estimation of grain color on
the F
2
and
F
2-3
plants, the genomic DNA of 241
recessive lines were selected and used for molecular
mapping. In all, 40 pairs of wheat SSR primers on
chromosome 3D were used to screen on the two
parents Chuanmai42 and CW565 of mapping populat-
ion. Primer sequences were described by Roder et al.
(1998), Pestsova et al. (2000), Daryl et al. (2004) and
Song et al. (2005), and then synthesized by Shanghai
Sangon Biological Engineering Technology & Services
Co. Ltd (http: // www.sangon.com).
The PCR reaction was performed in a volume of 15 µL
in a PTC100 Peltier Thermal Cycler. The reaction