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利用萝卜Ogura-CMS保持系和恢复系育性恢复基因Rfo的序列,开发了基于高通量竞争性等位基因特异性PCR(KASP)分型技术的SNP标记:Rfo-SNP1。利用该标记对24个萝卜品种的304个单株进行了育性恢复基因Rfo的基因型鉴定,其中289株获得基因分型结果,占整个群体的95.07%。从分型结果看,育性恢复基因Rfo的基因型为rfrf的有110株(占36.18%),基因型为Rf Rf和Rfrf的分别为156株(占51.32%)和23株(占7.57%)。在12个品种中获得基因型为rfrf的单株,可作为保持系并用于不育系转育。利用PCR-RFLP标记和回交群体单株育性鉴定,进一步证实Rfo-SNP1标记不但能够实现高通量检测,而且能够实现萝卜Ogura-CMS育性恢复基因Rfo基因型的精准鉴定。目前利用该方法已成功转育了6套稳定遗传的雄性不育系及保持系。
Using the sequence of the radish Ogura-CMS maintainer and restoring the fertility restorer Rfo, a SNP marker Rfo-SNP1 based on a high-throughput competitive allele-specific PCR (KASP) typing technique was developed. The genotypes of fertility restorer (Rfo) genotypes were detected in 304 individuals of 24 radish cultivars by using this marker, of which 289 were genotyped, accounting for 95.07% of the entire population. According to the typing results, 110 (36.18%) were genotypes of fertility restorer Rfrf and 156 (51.32%) and 23 (7.57%) were genotype Rff and Rfrf, respectively ). In 12 varieties obtained genotype rfrf single plant, can be used as a maintainer and used in sterile line. The results of PCR-RFLP and single-fertility identification of backcross showed that Rfo-SNP1 could not only detect high-throughput but also identify Rfo genotype of Ogura-CMS fertility restorer in radish. At present, 6 stable male sterile lines and maintainer lines have been successfully transferred by this method.