Variation of High-Molecular-Weight Glutenin Subunits and Gliadin in T.aestivum ssp. macha
In order to exploit new genetic resources and provide fundamental
materials for the breeding improvement of bread wheat quality, genetic
variation of high-molecular-weight glutenin subunits and gliadin in 29
macha wheat accessions were observed by acidic polyacrylamide-gel
electrophoresis and sodium dodecyl sulphate polyacrylamide-gel electrophoresis.
Nine HMW-glutenin subunits alleles and 9 combinations were identified.
Subunit null (82.8%), 7+8 (53.3%) and 2+12 (82.8%) scored the highest
frequency at Glu-A1, Glu-B1 and Glu-D1 loci, respectively.
In addition, subunits 7+9 (23.3%) was also found at higher frequency.
A total of 49 gliadin bands and 28 patterns were detected and the polymorph
among all accessions was identified in most of bands (97.96%). Furthermore,
all materials could be clustered into three major groups based on genetic
similarity coefficient. The results indicated that the variations of gliadin
among macha accessions were not associated with their geographic
In the endosperm of wheat (Triticum aestivum L.), the main
storage protein classes are glutenin and gliadin. The wheat baking quality
is controlled by the content and composition of wheat endosperm proteins.
Specific HMW-glutenin subunits have been closely associated with bread
making quality (Payne, 1987; He et al., 1999; Barro et al.,
2003; Edwards et al., 2007; Lefebvre and Mahmoudi, 2007). Glutenin
and gliadin are generally considered to contribute to the viscosity and
extensibility of gluten. Glutenins were composed of high-molecular-weight
(HMW) subunits and low-molecular-weight (LMW) subunits. HMW-glutenin subunits
are controlled by three gene loci, located on the long arm of chromosomes
1A, 1B and 1D and identified as Glu-A1, Glu-B1 and Glu-D1
(Payne, 1987). Gliadins on the other hand are encoded by the complex gene
family, which located on the short arms of group 1 and 6 chromosomes (Lafiandra
et al., 1984; Hassani et al., 2007).
In order to improve wheat quality breeding, it is essential to exploit
the valuable gene resource from wheat relative species. One of the oldest
hexaploid cultivated species, T. aestivum ssp. macha
(2n = 6x = 42, AABBDD), that was grown mainly in the west of Gelugia in
the past. It possessed stronger tillering ability, resist smut and moisture
tolerance. Because it has the same genome structure as the common wheat,
the useful genes of T. aestivum ssp. macha could
be easy transferred into common wheat. Datiashvili (1983) found that there
was better ratio between gliadin and glutenin in T. aestivum
ssp. macha. Fang et al. (1997) suggested that moisture
tolerance of macha wheat was controlled by a pair of dominant gene
and further scored a SSR marker Xgwm148 (Fang and Cai, 2003). Zhang et
al. (2003) further demonstrated that these genes are located on 5A
chromosome based on RAPD marker. Xiong et al. (2006) also reported
that macha wheat had higher plant height, strong tillering ability and
more spikelets. However, little information for macha was investigated.
This confined the utility of T. aestivum ssp. macha
in wheat improvement breeding. In the present study, were report on genetic
diversity of HMW-glutenin subunits and gliadin as evaluated in 29 T.
aestivum ssp. macha accessions from 11 countries.
MATERIALS AND METHODS
Plant Materials: A total of 29 accessions from different countries
were kindly provided by Dr. Harold Bockelaman, USDA-ARS, National Small Grains
Collection (Table 1). This study was conducted in 2007. Electrophoresis
was performed at the Triticeae Research Institute, Sichuan Agriculture University.
A-PAGE: According the standard protocol of ISTA acidic
polyacrylamide-gel electrophoresis (pH 3.1) (A-PAGE) (Draper, 1987), the
gliadin electrophoresis was carried out. Common wheat cultivar Chinese
Spring was used as reference.
SDS-PAGE: According to the procedures of Ng and Bushuk
(1987), the HMW-glutenin subunits were separated by sodium dodecyl sulphate
polyacrylamide-gel electrophoresis (SDS-PAGE). HMW-glutenin subunits were
identified according to Payne and Lawrence (1983). Common wheat cultivar
Chinese Spring (null/7+8/2+12) and Chuanyu12 (1/7+8/5+10) were used as
Statistic Analysis: The presence or absence of gliadin bands
was carefully recorded. The present bands were recorded as 1 and the bands absent
as 0. Based on the gliadin bands data, the Genetic Similarity (GS) was calculated
according to Nei`s (1973) method as GS = 2Nij/(Ni+ Nj),
where Ni is the number of the band in the material i; Nj
is the number of the gliadin band in the material j; Nij is the number
of the gliadin band shared by material i and j.
||A list of accessions and their country of origin
Using the unweighted pair-group method, a clustering analysis was carried out
based on GS index under NTSYS-pc (version 2.1) software.
Gliadin Variations: A total of 49 gliadin polymorphic
bands and 28 patterns were detected in 29 T. aestivum ssp.
macha accessions. The number of bands ranged from 19 to 34 per accession,
with the mean of 25. Ten bands were detected in α-, β- and γ-region,
respectively and 19 bands in ω-region. Twenty-two, nineteen, twenty-two
and twenty-six gliadin patterns were identified in α-, β-, γ-
and ω-region, respectively (Fig. 1). The genetic
similarity (GS) among T. aestivum ssp. macha ranged
from 0.29 to 0.95, with a mean of 0.61. Based on the gliadin GS coefficient,
the genetic relationships among macha wheat accessions were further
investigated by UPGMA program. The result indicated that all accessions
could be divided into three groups at the 0.58 level of GS (Fig.
2). Group 1 contained 19 accessions, including all accessions from
Hungary, Iran, Italy, United Kingdom and most of accessions from the Former
Soviet Union and Georgia. It could be further divided into two subgroups
at the 0.63 level of GS. Group 3 only contained one accessions (PI355513)
from Former Soviet Union. Group 2 contained 9 accessions, including 5
accessions from Switzerland, United States, Denmark, Sweden and Russian
Federation, respectively and 4 accessions from Georgia. Group 2 also could
be further divided into two subgroups. One accession (PI572909) from Georgia
formed subgroup 2, whereas other three accessions from Georgia were clustered
in another subgroup. Therefore, the clustering of macha wheat is
not associated with their geographic origin. These results also indicate
that the macha wheat accessions from Former Soviet Union and Georgia
had higher genetic variation than material from other sources.
||Gliadin patterns for 17 accessions of macha wheat in
APAGE profile Note:The name of material was listed in Table
1, CS: Chinese spring, α, β, γ and ω refer to the
corresponding gliadin region
||A dendrogram generated from gliadin
||HMW-GS of macha wheat accessions after SDS-PAGE Note:The
name of material was listed in Table 1, CK1:
Chinese spring, CK2: Chuanyu12
HMW-GS Glutenin Variations: A total of 9 HMW-GS subunits were
identified in all accessions, including 2, 5 and 2 subunits at Glu-A1,
Glu-B1and Glu-D1 loci, respectively (Table 2)
(Fig. 3). At the Glu-A1 locus, subunit null had the
highest frequency (82.8%), whereas subunit 1 had low frequency (17.2%). At Glu-B1
locus, subunits 7+8 was the predominant frequency 55.2%, followed by subunit
7+9 (24.1%) and the subunits 8, 20 and 6+8 with low frequency were also detected.
At Glu-D1 locus, only subunits 2+12 and 5+10 were identified and the
former subunit had the highest frequency (82.8%). Nine HMW-GS patterns were
identified among all accessions. The distribution of HMW-glutenin patterns are
listed in Table 3.
||The distribution frequency of HMW-GS alleles in macha
||The combination and distribution frequency of HMW-GS in macha
The pattern null/7+8/2+12 was detected in 10 accessions and had the highest
frequency. Four patterns, including null/7+9/2+12, null/7+8/5+10, 1/7+8/2+12
and 1/7+9/2+12, had higher frequency. The HMW-GS pattern 1/6+8/2+12 and null/20/2+12
were only observed at low frequency.
The gliadin patterns with a great diversity of variations have been
used as fingerprints, genotype identification, quality markers and so
on (Yan et al., 1992; Liu et al., 1999; Lang et al.,
2001). Zhang et al. (1995) suggested that A-PAGE could provide
an available method for identifying wheat germplasm. In this study, the
higher genetic diversity of gliadin was found, a total of 28 gliadin patterns
were identified in 29 T. aestivum ssp. macha. This
indicates that gliadin could provide a valuable and exact diversity marker
for T. aestivum ssp. macha accessions. These data
is supported by the results of Khachidze (1989) who also observed a great
diversity of variations in T. aestivum ssp. macha
and found some bands in the patterns that is species specific. Yakobashvili
and Naskidashvili (1988) reported that the greatest polymorphism (11 variants
of gliadin blocks) was found for the gliadin locus on chromosome 6A. One
of its variants was identical to block Gld 6A20 of bread wheat.
Six variants of blocks associated with chromosome 1B, 3 were identical
to blocks in bread wheat (Gld-1B4, Gld-1B8 and Gld-1B10).
Metakovsky and Iakobashvili (1990) further reported an additional component
mapped between Glu-B1 and Gli1B in the gliadin spectrum
of T. aestivum ssp. macha. The analyses of seed storage
protein components of the gliadin and glutenin in T. aestivum
ssp. compactum, T. aestivum ssp. sphaerococcum,
T. aestivum ssp. macha, T. aestivum ssp.
spelta had revealed limited variation at the tightly linked coding
loci Gli-D1/Glu-D3 and Glu-D1, located respectively
on the short and long arm of chromosome 1D and at the Gli-D2 locus,
positioned on the short arm of chromosome 6D (Lafiandra et al.,
In this study, a great diversity of variations on gliadin loci in T.
aestivum ssp. macha accessions, while the relationship between
genetic variety and other properties traits need further study. Meanwhile,
two accessions with the same gliadin bands had no diversity in all properties,
but had further close relative relations, such as PI352466 and PI355508
used in this study. So the genetic varieties between accessions need other
methods, like by molecular marker technology to detect and analyses.
In the tree of the cluster analysis, accessions from Georgia were clustered
together, but only few accessions were grouped with other counties accessions.
From the clustering dendrogram, the Georgia accessions could be divided into
two groups. Accessions PI572908, PI572913, PI572906 and PI572909 formed a group,
while others were another group. This indicates that there were two allelic
variety types on gliadin locus in Georgia accessions. In this study, the clustering
analysis indicated that the gliadin variations among T. aestivum ssp.
macha are not associated with their geographic origin. High level of HMW-GS
polymorphism was observed in the 29 T. aestivum ssp. macha
by SDS-PAGE method. The main HMW-glutenin patterns was null (Glu-A1),
In all ten Georgia accessions, the pattern (null, 7+8 and 2+12) had the highest
frequency, the pattern (null, 7+9, 2+12) had the similar performance in Former
Soviet Union accessions. The consistent efforts have been made to improve wheat
quality by accumulating good HMW-glutenin subunits, such as 1, 5+10. Yakobashvili`s
et al. (1988) indicated that the species included T. aestivum
ssp. macha, T. aestivum ssp. spelta and
T. aestivum ssp. aestivum were closely related through
comparison with HMW glutenins of 3 species by SDS-PAGE. The greatest polymorphism
in T. aestivum ssp. macha and T. aestivum ssp.
spelta was for Glu-1B, for which the greatest difference between
them was also found suggesting that they may had different donors of genome
This research was supported by the National Natural Science Foundation
of China (No. 30300219 and 30571163) and A Foundation for the Author of
National Excellent Doctoral Dissertation of PR China (No. 200357 and 200458).
Y.-L. Zheng was supported by the Program for Changjiang Scholars and Innovative
Research Teams in University of China (IRT0453).
Barro, F., P. Barcelo, P.A. Lazzeri, P.R. Shewry, J. Ballesteros and N.A. Martin, 2003. Functional properties of flours from field grown transgenic wheat lines expressing the HMW glutenin subunit 1AX1 and 1DX5 genes. Mol. Breeding, 12: 223-229.
CrossRef | Direct Link |
Datiashvili, N.A., 1983. Protein content in some source species and experimentally produced forms of wheat endemic in the Georgian SSR. Bull. Aca. Sci. Georgian SSR, 110: 121-124.
Draper, S.R., 1987. ISTA and Variety committee: Report of the working group for biochemical tests for cultivar identification 1983-1986. Seed Sci. Tech., 15: 431-434.
Direct Link |
Edwards, N.M., M.C. Gianibelli T.N. McCaig, J.M. Clarke, N.P. Ames, O.R. Larroque and J.E. Dexter, 2007. Relationships between dough strength, polymeric protein quantity and composition for diverse durum wheat genotypes. J. Cereal Sci., 45: 140-149.
Fang, X.W. and S.B. Cai, 2003. SSR marker and location of gene controlling waterlogging resistance in macha wheat. Jiangsu J. Agric. Sci., 19: 253-254.
Hassani, M.E., M.R. Shariflou, M.C. Gianibelli and P.J. Sharp, 2008. Characterisation of a ω-gliadin gene in Triticum tauschii. J. Cereal Sci., 47: 59-67.
He, G.Y., L. Rooke, S. Steele, F. Bekes and P. Gras et al., 1999. Transformation of pasta wheat (Triticum turgidum L. var. durum) with high-molecular-weight glutenin subunit genes and modification of dough functionality. Mol. Breeding, 5: 377-386.
Hua, L., W. Yusheng and Z. Hui, 1999. Preliminary construction and application of gliadin fingerprints database of Chinese wheat germplasm. Acta Agron. Sin., 25: 674-682.
Direct Link |
Khachidze, T.O., 1989. Electrophoretic banding patterns of the gliadin proteins in grains of Georgian endemic wheats. Soobshcheniya Akademii Nauk Gruzinskoi SSR, 134: 625-628.
Lafiandra, D., D.D. Kasarda and R. Morris, 1984. Chromosomal assignment of genes coding for the wheat gliadin protein components of the cultivars Cheyenne and Chinese Spring by two dimensional (two-pH) electrophoresis. Theor. Applied Genet., 68: 531-539.
Lafiandra, D., S. Masci, R. Dovidio, O.A. Tanzarella, E. Porceddu and B. Margiotta, 1992. Relationship between the D genome of hexaploid wheats (AABBDD) and Ae. Squarrosa as deduced by seed storage proteins and molecular marker analyses. Here. Landskrona, 116: 233-238.
Lang, M.L., S.Y. Lu and R.Z. Zhang, 2001. Analysis of the genetic evolution of gliadin composition in the major wheat cultivars grown in North China. Acta Agron. Sin., 27: 958-966.
Lefebvre, J. and N. Mahmoudi, 2007. The pattern of the linear viscoelastic behaviour of wheat flour dough as delineated from the effects of water content and high molecular weight glutenin subunits composition. J. Cereal Sci., 45: 49-58.
CrossRef | Direct Link |
Metakovsky, E.V. and Z.A. Iakobashvili, 1990. Homology of chromosomes of Triticum macha Dek. et Men. and T. aestivum L. as shown with the help of genetic makers. Genome, 33: 755-757.
Nei, M., 1973. Analysis of gene diversity in subdivided populations. Proc. Natl. Acad. Sci. USA., 70: 3321-3323.
PubMed | Direct Link |
Ng, P.K.W. and W. Bushuk, 1987. Glutenin of Marquis wheat as a reference for estimating molecular weights of glutenin subunits by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cereal Chem., 64: 324-327.
Direct Link |
Payne, P.I. and G.J. Lawrence, 1983. Catalogue of alleles for the complex gene loci, Glu-A1, Glu-B1 and Glu-D1 which code for high molecular-weight subunits of glutenin in hexaploid wheat. Cereal Res. Commun., 11: 29-35.
Payne, P.I., 1987. Genetics of wheat storage proteins and the effect of allelic variation on breadmaking quality. Ann. Rev. Plant Physiol., 38: 141-153.
Direct Link |
Xianwen, F., C. Yang and C. Shibin, 1997. Genetic evaluation of waterlogging tolerance in Triticum macha. Jiangsu J. Agric. Sci., 13: 73-75.
Direct Link |
Xiong, L.J., W. Li and Y.L. Zheng, 2006. Analysis in principle agronomic traits of Triticum macha Dekaprel et Menabde. Chin. Agric. Sci. Bull., 22: 118-122.
Yakobashvili, Z.A. and P.P. Naskidashvili, 1988. SDS-PAGE study of the polymorphism of high-molecular-weight (HMW) glutenin in hexaploid wheats of the species Triticum macha, T. spelta and T. vavilovii. Soobshcheniya Akademii Nauk Gruzinskoi SSR., 130: 397-400.
Yakobashvili, Z.A., P.P. Naskidashvili and E.V. Metakovskii, 1988. Study of gliadin polymorphism in Triticum macha Dek. et Men. Soobshcheniya Akademii Nauk Gruzinskoi SSR., 130: 609-612.
Yan, Q.C., Y.J. Huang and Y. Xu, 1992. Cultivar identification of barley and wheat with standard reference method from International Seed Testing Association (ISTA). Acta Agron. Sin., 18: 61-68.
Zhang, J.L., S.B. Cai, G.X. Zhang, J.B. Wei, C.Q. Zhang and Z.Q. Ma, 2003. Genetic mapping of genes conferring waterlogging-tolerance in Triticum macha using RAPD markers. J. Nanjing Agric. Uni., 26: 7-10.
Direct Link |
Zhang, X., X. Yang and Y.S. Dong, 1995. Genetic analysis of wheat germplasm by acid polyacrylamide gel electrophoresis of gliadins. Sci. Agric. Sin. A, 28: 25-32.
Direct Link |