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油菜抗咪唑啉酮类除草剂基因BnALS1R是从抗性突变体M9中克隆获得,抗性基因BnALS1R与野生型基因BnALS1存在1处SNP,即乙酰乳酸合酶第638位丝氨酸残基被天冬酰胺酸替代。为获得油菜抗除草剂基因BnALS1R的分子标记,根据该处点突变,结合获得的BnALS3与BnALS1序列,开发30条等位基因特异PCR(allele-specific PCR,AS-PCR)引物,采用筛选出的3条AS-PCR引物在F2、BC1和BC2群体中进行PCR扩增。结果表明,该标记有效检测出群体中存在的3种基因型,其分离比分别为1∶2∶1、1∶1、1∶1,均遵循单基因遗传规律。应用该标记对获得的抗性恢复系进行PCR扩增,结果发现所有抗性恢复系均能扩增出抗性基因BnALS1R目的条带,表明3条标记引物可应用于抗性基因的检测。AS-PCR标记的获得将促进以抗性基因进行油菜抗除草剂分子标记辅助选择育种。
The resistance gene BnALS1R of rapeseed imidazolinone herbicide gene BnALS1R was cloned from the resistant mutant M9. One SNP exists between the resistance gene BnALS1R and the wild type BnALS1, that is, the serine residue at 638 of acetolactate synthase is replaced by asparagine Acid replacement. In order to obtain the molecular markers of the herbicide resistance gene BnALS1R, 30 allele-specific PCR (AS-PCR) primers were developed based on the point mutation and BnALS3 and BnALS1 sequences obtained. Three AS-PCR primers were PCR amplified in F2, BC1 and BC2 populations. The results showed that the marker could effectively detect the three genotypes in the population, with the segregation ratios of 1: 2: 1, 1: 1 and 1: 1, respectively, all following the single-gene inheritance pattern. The marker was used to amplify the resistant restorer lines. All of the restorer lines could amplify the BnALS1R-resistant bands, indicating that three marker primers could be used to detect the resistance genes. Obtaining of AS-PCR markers will promote rapeseed-resistant herbicide molecular marker-assisted selection breeding with resistance genes.