强筋小麦分子标记多重PCR体系的构建与应用

Establishment and Application of Multiplex PCR System Based on Molecular Markers of Glutenin Subunit Genes (Loci) Related to Strong-gluten in Wheat

面筋强度与小麦高低分子量谷蛋白亚基种类(组合)密切相关。以12个已知亚基基因组成的品种为对照,选用基因(位点)Axnull、Bx7OE、Dx5、Glu-A3d、Glu-B3i和Glu-B3的标记,构建多重PCR体系。以该体系检测对照的结果与已知基因型完全一致,一次PCR可同时间接检测7个与强筋有关基因(位点)Ax1/Ax2*、Bx7OE、Dx5、Glu-A3d、Glu-B3i和Glu-B3。对62个陕西小麦品种的检测结果表明,优质强筋亚基基因(位点)Ax1/Ax2*、Dx5、Glu-A3d、Glu-B3i和Glu-B3的比例依次为56.5%、9.6%、33.9%、1.6%和64.4%,所有品种均不含Bx7OE,携带0、1、2和3个及以上基因(位点)的品种分别占6.5%、33.9%、48.3%和11.3%。说明聚合多个强筋亚基基因(位点)的品种频率较低,通过优质亚基基因的聚合育种,可望改善陕西小麦品种的面筋品质。本研究所构建的强筋小麦分子标记检测的多重PCR体系检测结果稳定且可靠,可用于小麦种质资源的快速评价和强筋小麦分子辅助选育。

Wheat strong-gluten quality is closely correlated with combinations of high-molecular-weight glutenin subunits （HMW-GS） and low-molecular-weight glutenin subunits （LMW-GS）. The multiplex PCR system is a rapid and efficient approach to evaluate wheat germplasm and its quality in wheat. In this study, we developed a multiplex PCR system confering molecular markers on Ax1/Ax2＊, Bx7OE, Dx5, Glu-A3d, Glu-B3i genes, and Glu-B3 locus and validated it with 12 cultivars with known subunit at each locus. This multiplex PCR system was proved to be effective and stable to amplify target bands for these genes （locus）, and used to evaluate the glutenin subunit genes （locus） accociated with strong-gluten in 62 major cultivars in wheat production in Shaanxi province, China. The results showed that the frequencies of genes Ax1/Ax2＊, Dx5, Glu-A3d, Glu-B3i, and locus Glu-B3 in the 62 cultivars were 56.5%, 9.6%, 33.9%, 1.6%, and 64.4%, respectively, whereas gene Bx7OE was not found. Most of the cultivars carried two-gene （locus） combinations with the frequency of 48.3%, a few cultivars carried a single gene or locus （33.9%）. The frequency of cultivars carrying three or four-gene （locus） combinations was 11.3%. The remaining cultivars （6.5%） were free of above elite gene （locus）. Therefore, the frequency of combination of multiple strong-gluten subunits gene （locus） was low in cultivars from Shaanxi Province, which could be promoted through germplasm introduction and traditional breeding aided by molecular marker selection. The multiplex PCR system developed in this study may serve as a rapid and efficient method to select materials pyrimiding multiple genes （loci） associated with strong-gluten in wheat breeding for quality.