簇毛麦6VS特异转录序列P21461及P33259的获得及其分子标记在鉴定小麦-簇毛麦抗白粉病育种材料中的应用

Developing of Specific Transcription Sequences P21461 and P33259 on Dasypyrum villosum 6VS and Application of Molecular Markers in Identifying Wheat–D. villosum Breeding Materials with Powdery Mildew Resistance

簇毛麦6V#2S和6V#4S染色体臂分别携带抗白粉病基因Pm21和Pm V,在与小麦的杂种后代中,抗病基因与外源染色体臂共分离。开发鉴定2条外源染色体臂间多态性的序列,尤其是遗传信息相对缺乏的6V#4S染色体臂的序列,对于其在遗传与育种上的应用具有重要意义。本研究以携带6V#4S·6DL染色体的小麦易位系Pm97033及感病小麦亲本宛7107接种白粉菌的叶片转录组数据为资源,通过差异基因筛选、共线性分析、簇毛麦基因组扩增及测序验证的方法,鉴定出来自6V#4S的表达序列P21461和P33259,其中基于P21461序列设计的引物P461-5在簇毛麦6V#2S和6V#4S染色体臂的扩增产物具有30 bp的In Del和4 nt的多态性。用该引物转化的标记P461-5a可以鉴定抗白粉病小麦品种和高代品系所含的外源染色体,显示其在簇毛麦抗源鉴别和小麦抗病育种辅助选择中潜在的应用价值。根据P33259开发的标记P259-1可以对含有6V#4S染色体臂的材料进行特异扩增,但对6V#2S·6AL易位染色体没有扩增产物,因此P259-1可作为6V#4S·6DL易位染色体的特异分子标记。q RT-PCR分析结果显示,P21461的表达不受白粉菌诱导,而P33259在接菌后12 h和24 h的转录水平比接菌前提高约2倍,推测其可能参与Pm97033与白粉菌的早期互作。

Dasypyrum villosum carries powdery mildew （PM） resistance gene Pm21 and PmV on its chromosome arms of 6V#2S and 6V#4S,respectively. The two resistant genes are co-segregated with exogenous chromosome arms in the offspring of hybrids between translocation lines andcommon wheat varieties. Developing polymorphic expression sequences to identify the two exogenous chromosomal arms is of great significance fortheir application in genetics and breeding, especially for 6V#4S chromosome, on which hereditary informations relatively lack. In this study,transcriptome of leaves of 6V#4S·6DL translocation line Pm97033 and common wheat line Wan7107 inoculated with Bgt was used as resources forsequences screening. As results, two unigenes of P21461 and P33259 from 6V#4S were identified by differential gene screening, synteny analysis,genomic DNA amplification and products sequencing of D. villosum accordingly. By using a pair of designed primer P461-5 based on P21461 sequence,it was found that there were 30 bp of InDel and 4 nt polymorphism between chromosomal arms of 6V#2S and 6V#4S. By the newly developed markerP461-5a, which retains the InDel polymorphism and truncates the product size amplified by P461-5,