大豆CMS-RN型不育系育性恢复基因GmRf1的初步鉴定及其分子标记开发

Preliminary Identification and Molecular Marker Development of the Restorer-of-fertility Gene GmRf1 of CMS-RN Type Sterile Lines in Soybean

在大豆杂种优势利用中,主要基于三系法进行杂交大豆品种选育。恢复系作为杂交种的父本,其所含的育性恢复(Rf,restorer-of-fertility)基因起决定作用。前期对大豆RN型细胞质雄性不育恢复系的育性恢复基因GmRf1进行了精细定位。本研究在此基础上,对GmRf1定位区间内的候选基因进行功能注释、亚细胞定位预测、序列比对和差异表达分析,明确了Glyma.16G161900基因在恢复系JLR230中所编码的1个576个氨基酸的PPR(Pentatricopeptide repeats)蛋白为GmRf1。进一步对含有GmRf1的对照材料Williams82与母本不育系JLCMS204A进行测交及F_(1)植株花粉育性鉴定,验证了GmRf1可以恢复CMS-RN型不育系育性。最后利用GmRf1在亲本间存在的单核苷酸突变位点,开发了功能性分子标记Rf1-dCAPS-2和Rf1-dCAPS-3。上述研究将为今后通过分子标记筛选或辅助选育含GmRf1基因型材料,以及通过基因工程手段创制新型恢复系奠定了理论和技术基础。

The breeding of hybrid soybean varieties is mainly based on the three-line method(based on cytoplasmic male sterility).As the male parent of the hybrid,the restorer-of-fertility(Rf)gene in the restorer line plays a decisive role.In the former study,we mapped the Rf gene GmRf1 for the soybean RN type cytoplasmic male sterile line.This study conducted functional annotation,subcellular location prediction,sequence alignment and differential expression analysis of candidate genes in the physical interval of GmRf1.We identified that the PPR(Pentatricopeptide repeats)gene Glyma.16G161900 encoding for 576 amino acids encoded in male restorer line JLR230 is GmRf1.By crossing Williams82 containing GmRf1 with the female sterile line JLCMS204A,the pollen fertility of F_(1) plants was identified demonstrating that GmRf1 could restore the fertility of CMS-RN sterile lines.We further developed functional molecular markers Rf1-dCAPS-2 and Rf1-dCAPS-3 based on the single nucleotide mutation sites of GmRf1 between female and male parents.Collectively,this study laid a theoretical and technical foundation for the future marker-assisted selection of GmRf1,as well as the creation of new restorer lines through genetic engineering.