具有成本效益的双等位基因竞争特异性聚合酶链反应标记，用于同源基因，以促进小麦的选育。
Cost-effective duplex Kompetitive Allele Specific PCR markers for homologous genes facilitating wheat breeding.
机构信息
CIMMYT-JAAS Joint Center for Wheat Diseases, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, Jiangsu, China.
Lianyungang Institute of Agricultural Sciences, Lianyungang, 222000, Jiangsu, China.
出版信息
BMC Plant Biol. 2023 Mar 1;23(1):119. doi: 10.1186/s12870-023-04116-y.
BACKGROUND
Owing to successful cloning of wheat functional genes in recent years, more traits can be selected by diagnostic markers, and consequently, effective molecular markers will be powerful tools in wheat breeding programs.
RESULTS
The present study proposed a cost-effective duplex Kompetitive Allele Specific PCR (dKASP) marker system that combined multiplex PCR and KASP™ technology to yield twice the efficiency at half the cost compared with the common KASP™ markers and provide great assistance in breeding selection. Three dKASP markers for the major genes controlling plant height (Rht-B1/Rht-D1), grain hardness (Pina-D1/Pinb-D1), and high-molecular-weight glutenin subunits (Glu-A1/Glu-D1) were successfully developed and applied in approved wheat varieties growing in the middle and lower reaches of the Yangtze River and advanced lines from our breeding program. Three markers were used to test six loci with high efficiency. In the approved wheat varieties, Rht-B1b was the most important dwarfing allele, and the number of accessions carrying Pinb-D1b was much greater than that of the accessions carrying Pina-D1b. Moreover, the number of accessions carrying favorable alleles for weak-gluten wheat (Null/Dx2) was much greater than that of the accessions carrying favorable alleles for strong-gluten wheat (Ax1 or Ax2/Dx5). In the advanced lines, Rht-B1b and Pinb-D1b showed a significant increase compared with the approved varieties, and the strong-gluten (Ax1 or Ax2/Dx5) and weak-gluten (Null/Dx2) types also increased.
CONCLUSION
A cost-effective dKASP marker system that combined multiplex PCR and KASP™ technology was proposed to achieve double the efficiency at half the cost compared with the common KASP™ markers. Three dKASP markers for the major genes controlling PH (Rht-B1/Rht-D1), GH (Pina-D1/Pinb-D1), and HMW-GS (Glu-A1/Glu-D1) were successfully developed, which would greatly improve the efficiency of marker-assisted selection of wheat.
背景
近年来，由于小麦功能基因的成功克隆，可以通过诊断标记物选择更多的性状，因此，有效的分子标记将成为小麦育种计划的有力工具。
结果
本研究提出了一种具有成本效益的双竞争等位基因特异性 PCR（dKASP）标记系统，该系统结合了多重 PCR 和 KASP™技术，与普通 KASP™标记相比，效率提高了一倍，成本降低了一半，为育种选择提供了很大的帮助。成功开发并应用了三个控制株高的主基因（Rht-B1/Rht-D1）、籽粒硬度（Pina-D1/Pinb-D1）和高分子量谷蛋白亚基（Glu-A1/Glu-D1）的 dKASP 标记，用于鉴定生长在长江中下游的审定小麦品种和本研究的育成品系。三个标记用于高效测试六个基因座。在审定品种中，Rht-B1b 是最重要的矮化等位基因，携带 Pinb-D1b 的品种数量远远大于携带 Pina-D1b 的品种数量。此外，携带弱筋小麦（Null/Dx2）有利等位基因的品种数量远远大于携带强筋小麦（Ax1 或 Ax2/Dx5）有利等位基因的品种数量。在育成品系中，Rht-B1b 和 Pinb-D1b 与审定品种相比有显著增加，强筋（Ax1 或 Ax2/Dx5）和弱筋（Null/Dx2）类型也有所增加。
结论
提出了一种具有成本效益的 dKASP 标记系统，该系统结合了多重 PCR 和 KASP™技术，与普通 KASP™标记相比，效率提高了一倍，成本降低了一半。成功开发了三个控制 PH（Rht-B1/Rht-D1）、GH（Pina-D1/Pinb-D1）和 HMW-GS（Glu-A1/Glu-D1）的主基因的 dKASP 标记，这将极大地提高小麦分子标记辅助选择的效率。
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