基于RNA-seq数据的小麦条锈菌SSR标记开发

Development of SSR Markers Based Puccinia striiformis f. sp. tritici Transcriptome

为开发小麦条锈菌基因组编码区内多态性高的SSR标记,应用MISA软件从4个小麦条锈菌分离物转录组测序数据中搜索SSR位点并设计相应引物。结果表明,在小麦条锈菌转录组2 941条unigenes中发现6 083个SSR位点,SSR发生频率为14.13%,平均每9.84 kb出现1个SSR位点。获得的SSR有144种重复基序,单核苷酸和三核苷酸是主要重复类型,分别占SSR总数的42.33%和39.90%。T和AAC、ATC、CAT分别是单核苷酸和三核苷酸的优势重复基序。转录组中SSR重复次数以5~12次为主,基序长度主要集中在12~20 bp。随机合成100对SSR引物对3个小麦条锈菌分离物基因组DNA进行PCR扩增,有87对引物可以扩增出条带。总之,这些基于转录组发掘的SSR位点出现频率高、类型丰富,在小麦条锈菌群体遗传结构研究、分子标记开发等方面具有一定的应用潜力。

In order to develop genetic simple sequence repeat（ SSR） marker from a unigene set assembled from Puccinia striiformis f. sp. tritici（ Pst） transcriptomic reads,the SSR information in four Pst transcriptomes was analyzed by MISA software and primers were designed. A total of 6 083 SSR loci were found in 2 941 unigenes in this study. The frequency of SSR was 14. 13% and the density of distribution was 9. 84 kb per SSR. Among the SSRs in Pst transcriptome,144 kinds of repeat motifs were found. Single nucleotide and trinucleotide were the major repeated types,accounting for 42. 33% and 39. 90%,respectively. T and AAC/ATC/CAT were the superior motif types in single and trinucleotide repeat motifs. Most of the repeat number of SSRs was from 5 to 12,and the length of the motifs ranged from 12 to20 bp. A total of 100 primer pairs were designed randomly from the SSR loci and 87 pairs were verified with genomic DNA of 3 Pst isolates by PCR. These SSR loci in Pst transcriptome,with high frequency and rich type,will be useful for studies of population genetics and molecular markers in Pst.