Development of SSR Markers Associated with Polima CMS Fertility Restorer in Brassica napus L. and Its Application

Abstract

The Polima system of cytoplasmic male sterility (Pol CMS) was widely used for hybrid seed production, but with high probability of occurrence of maternal false hybrid due to the fertility of Polima CMS (Pol CMS) line greatly affected by the ambient temperature, therefore, the detection of maternal false hybrids is the key to purity identification. In present study, the SSR markers were designed based on the flanking sequences of Pol CMS restorer gene Rfp , and application of SSR markers in hybrid seed purity test of rapeseed. The main results were as follows: the sequences of Rfp were subjected to basic local alignment search tool queries against the Brassica rapa genome to determine chromosome positions, and 32 SSR markers were developed based on the upstream and downstream sequences of Rfp . Among the 32 SSR markers, one codominant marker was polymorphic between Pol CMS sterile and restorer line, the PCR product length was 162 bp in Pol CMS sterile lines, and 184 bp in restorer lines. Based on the PCR sequence, two insert fragments (15 bp and 7 bp) were identified in Pol CMS restorer lines. The SSR marker co-segregated with plant fertility of individuals in the “fengyou737” F 2 population. The purity of “fengyou737”, “fengyou320” and “fengyou958” were identified by means of PCR-SSR markers and field-planting, and the results of two ways were highly consistent, indicated that the codominant SSR marker tightly linked to Rfp , and the marker can detect paternal and maternal false hybrids. The present findings will provide technical assistance for hybrid seed purity test and genetic diversity analysis in rapeseed.