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利用SSRIT软件对NCBI上蝴蝶兰的8 188条EST序列按照碱基重复单元数8以上的标准进行SSR位点查找,发掘出246条EST序列,共含有261个SSR位点,检出率为3.19%。二核苷酸重复是最主要的重复类型,占SSR总数的94.25%。SSR的不同重复单元中,最常见的重复单元是:(TA)n和(GA)n,其余出现频率较高的依次为(TC)n、(AT)n和(AG)n。设计了32对EST-SSR引物,以蝴蝶兰品种‘V31’的DNA为模板进行PCR扩增,发现16对引物能扩增出预期产物。进一步用这16对引物对16个蝴蝶兰品种(系)进行PCR扩增,9对引物有多态性,多态性条带数在2~12个之间。结果表明,设计开发的蝴蝶兰的EST-SSR标记是有效的。
Using SSRIT software, 8 188 EST sequences of Phalaenopsis on NCBI were searched for SSR loci according to the number of base repeat units of 8 or more, and 246 EST sequences were found, which contained 261 SSR loci with a detection rate of 3.19 %. Dinucleotide repeats were the most common repeat types, accounting for 94.25% of the total number of SSRs. Among the different SSR repeat units, the most common repeat units are: (TA) n and (GA) n, and the other most frequently occurring recurring units are (TC) n, (AT) n and (AG) n. Thirty-two pairs of EST-SSR primers were designed and PCR amplified using the DNA of Phalaenopsis species ’V31’ as a template. Sixteen pairs of primers were designed to amplify the expected product. Sixteen pairs of primers were further used to amplify the 16 Phalaenopsis cultivars (lines). Nine pairs of primers were polymorphic and the number of polymorphic bands was between 2 and 12. The results showed that EST-SSR markers of phalaenopsis were effective.