白菜EST-SSR信息分析与标记的建立

Data Mining for SSRs in ESTs and EST-SSR Marker Development in Chinese Cabbage

数量迅速增加的EST为开发新的SSR标记提供了宝贵的资源。本研究对4584条白菜EST进行了搜索,共检索出474个SSR,检出率为10·3%,包括40种重复基元。其中二核苷酸和三核苷酸重复单元的EST-SSR占主导地位,二者出现的频率基本相近,占总SSR的近83%;其它重复类型所占比例均不足5%。GA和GAA是二、三核苷酸中的优势重复类型,分别占二、三核苷酸重复类型的71·5%和37·5%。设计了15对EST-SSR引物,在合适的PCR反应体系下,以构建EST的白菜自交系A的DNA为模板,对设计的EST-SSR引物进行筛选,发现15对EST-SSR引物都能扩增出产物。进一步用这些可扩增的引物对28个白菜品种进行PCR扩增,发现7对引物显示多态性,占引物总数的46·7%。此结果表明,根据EST建立EST-SSR标记是一条简便而又有效的途径。

Expressed sequence tags （ESTs）, increased rapidly in number recently, are important resources for development of new SSR markers. In this study, 4 584 ESTs of Chinese cabbage were screened and 474 SSRs including 40 kinds of repeat motifs were mined out, accounting for 10. 3% of ESTs. Dinucleotide and trinucleotide repeats, with similar frequency and accounting for 83% together in all SSRs, were dominant, while the frequency for other repeat type is below 5% each. GA and GAA are the most frequent motifs, accounting fo 71.5% and 37.5% in dinucleotide and trinucleotide repeats respectively. 15 primer pairs for EST-SSRs were designed. Under a suitable PCR system, primers were screened against genomic DNA of inbred line A from which the cDNA library was constructed, and all the 15 primer pairs showed the amplification. Then all the primers available were subjected to PCR for DNAs from 28 Chinese cabbage varieties and 7 primer sets showed polymorphisms, accounting for 46.7% of total primers tested. Results indicate that it is a simple and effective approach to develop SSR markers based on ESTs.