花椰菜细胞质雄性不育基因特异PCR标记的筛选

Identification of PCR Markers Associtated with Cytoplasmic Male Sterility in Brassica oleracea var. Botrytis

基于同源序列的候选基因法(homology based candidate gene method),通过检索 NCBI核酸及蛋白数据库,获得细胞质雄性不育(cytoplasmic male sterility CMS)相关的基因或开放读码框。生物学软件分析,根据保守区设计5对特异引物,PCR扩增,其中引物P9/P10在花椰菜细胞质雄性不育系 knxd612 中特异扩增出 313 bp的片段。单株检测,RT PCR分析,斑点杂交鉴定,确定此片段为花椰菜细胞质雄性不育系 knxd612 所特有。序列分析表明该片段与Ogura型胞质不育萝卜,不育相关开放读码框orf138的同源性高达98%。初步结果显示实验所用不育花椰菜胞质亦可能为Ogura型,为进一步从分子水平研究花椰菜细胞质雄性不育打下了坚实的基础。

The homology-based candidate gene method was used to identify the specific PCR markers linked to cytoplasmic male sterility (CMS) in cauliflower(Brassica oleracea var. Botrytis.). By searching the DNA and protein (data-)base of NCBI , correlative genes or open reading frames were identified . Through analysis of biosoft, based on the conservative regions ,five primers were designed . Among them, only primer P9/P10 produced a 313- bp specific fragment. Identified by individual plant testing , analysis of RT-PCR and dot blot ,this fragment was only found in CMS cauliflower knxd612.Analysis of the sequence indicated it was high homologous(98%) with orf138 of Ogura CMS radish. Preliminary results suggested that the cytoplasmic type of CMS cauliflower knxd612 might belong to Ogura type. This research offered a good foundation to further investigate the CMS mechanism of cauliflower at molecular (level.)