Comparison of low molecular weight glutenin subunits identified by SDS-PAGE, 2-DE, MALDI-TOF-MS and PCR in common wheat

Springer Science and Business Media LLC - 2010
Li Liu1, Tatsuya Ikeda2, Gérard Branlard3, Roberto J. Peña4, William J. Rogers5, Silvia E. Lerner6, Maria de Los Angeles Kolman5, Xianchun Xia1, Linhai Wang1, Wujun Ma7, R. Appels7, Hisashi Yoshida8, Aili Wang9, Yueming Yan9, Zhonghu He1,10
1Institute of Crop Science, National Wheat Improvement Center/The National Key Facility for Crop Genetic Resources and Genetic Improvement, Chinese Academy of Agricultural Sciences (CAAS), China
2National Agriculture and Food Research Organization, Fukuyama, Japan
3INRA Station d'Amelioration des Plantes, Domaine de Crouelle, Clermont- Ferrand, France
4CIMMYT Mexico, Apdo, Postal, Mexico, Mexico
5CIISAS, CICPBA-BIOLAB AZUL, Facultad de Agronomía, Universidad Nacional del Centro de la Provincia de Buenos Aires, Azul, Provincia de Buenos Aires, Argentina. CONICET INBA -CEBB-MdP
6CRESCAA, Facultad de Agronomía, Universidad Nacional del Centro de la Provincia de Buenos Aires, Azul, Provincia de Buenos Aires, Argentina
7Western Australia Department of Agriculture and Food, State Agriculture Biotechnology Center, Murdoch University, Murdoch, Australia
8National Agriculture and Food Research Organization, Tsukuba, Japan
9Key Laboratory of Genetics and Biotechnology, College of Life Science, Capital Normal University, Beijing, China
10International Maize and Wheat Improvement Center (CIMMYT) China Office, Beijing, China

Tóm tắt

Abstract Background Low-molecular-weight glutenin subunits (LMW-GS) play a crucial role in determining end-use quality of common wheat by influencing the viscoelastic properties of dough. Four different methods - sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), two-dimensional gel electrophoresis (2-DE, IEF × SDS-PAGE), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and polymerase chain reaction (PCR), were used to characterize the LMW-GS composition in 103 cultivars from 12 countries. Results At the Glu-A3 locus, all seven alleles could be reliably identified by 2-DE and PCR. However, the alleles Glu-A3e and Glu-A3d could not be routinely distinguished from Glu-A3f and Glu-A3g, respectively, based on SDS-PAGE, and the allele Glu-A3a could not be differentiated from Glu-A3c by MALDI-TOF-MS. At the Glu-B3 locus, alleles Glu-B3a, Glu-B3b, Glu-B3c, Glu-B3g, Glu-B3h and Glu-B3j could be clearly identified by all four methods, whereas Glu-B3ab, Glu-B3ac, Glu-B3ad could only be identified by the 2-DE method. At the Glu-D3 locus, allelic identification was problematic for the electrophoresis based methods and PCR. MALDI-TOF-MS has the potential to reliably identify the Glu-D3 alleles. Conclusions PCR is the simplest, most accurate, lowest cost, and therefore recommended method for identification of Glu-A3 and Glu-B3 alleles in breeding programs. A combination of methods was required to identify certain alleles, and would be especially useful when characterizing new alleles. A standard set of 30 cultivars for use in future studies was chosen to represent all LMW-GS allelic variants in the collection. Among them, Chinese Spring, Opata 85, Seri 82 and Pavon 76 were recommended as a core set for use in SDS-PAGE gels. Glu-D3c and Glu-D3e are the same allele. Two new alleles, namely, Glu-D3m in cultivar Darius, and Glu-D3n in Fengmai 27, were identified by 2-DE. Utilization of the suggested standard cultivar set, seed of which is available from the CIMMYT and INRA Clermont-Ferrand germplasm collections, should also promote information sharing in the identification of individual LMW-GS and thus provide useful information for quality improvement in common wheat.

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