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利用集团分离分析法(Bulked segregant analysis BSA),以辣椒细胞质雄性不育系BU-12、恢复系RF-12为材料共筛选了336条RAPD引物,其中引物S418在恢复系中呈现特异性扩增,得到一条约3000bp的特异片段。回扩得到两条片段,测序表明大小为1515bp,1162bp。荧光原位杂交证实1515bp片段为恢复系特有,命名为S418_(1515)。序列分析表明S418_(1515)为一新发现的序列,Blastn序列比对同源性小于40%,tBlastx比对发现该序列与水稻2、4、7、10号染色体的几个BAC克隆上的序列高度同源。推测可能与其具有相似的编码功能,为进一步从分子水平研究辣椒育性恢复打下了坚实的基础。根据测序结果设计特异引物,将S418_(1515)转化成特异PCR标记,证明能用于候选材料的初筛。
A total of 336 RAPD primers were screened by bulked segregant analysis (BSA) using the pepper cytoplasmic male sterile line BU-12 and the restorer line RF-12. The primer S418 showed specific amplification in the restorer line , Get a specific fragment of about 3000bp. Two fragments were obtained by backcrossing, and sequencing showed that the size was 1515bp and 1162bp. Fluorescence in situ hybridization confirmed that the 1515bp fragment was unique to the restorer line and named S418_ (1515). Sequence analysis showed that S418_ (1515) was a newly discovered sequence with Blastn sequence homology less than 40%. The sequence of tBlastx was found to be similar to that of several BAC clones on chromosomes 2, 4, 7 and 10 Highly homologous. It is speculated that it may have a similar coding function, laying a solid foundation for further research on pepper fertility restoration at the molecular level. According to the sequencing results, specific primers were designed and S418_ (1515) was transformed into specific PCR markers, which can be used to screen candidate materials.