大麻SSR标记的开发及指纹图谱的构建

Development of SSR Markers and Construction of Fingerprint for Cannabis( Cannabis sativa L. )

表达序列标签(expressed sequence tags,EST)是开发SSR标记的重要资源。本研究从NCBI中大麻EST数据库中检索到1114个SSR,分布于989条EST序列中,占EST总数的7.66%。其中三、六核苷酸重复基元类型居多,分别占EST-SSR总数的39.84%和34.56%,统计得到三核苷酸重复类型47种,六核苷酸重复类型113种。利用部分EST-SSR序列设计49对SSR引物,其中40对引物有扩增产物,占所设计引物总数的81.63%。进一步用这些引物在24个大麻品种进行多态性检测,29对引物显示多态性,占可扩增引物的72.5%。利用E-7、E-8、E-11、E-18、E-21、E-39及E-48共7对引物构建了24份供试材料的SSR指纹图谱。本研究结果证明了基于大麻EST信息建立SSR标记是一种有效而又可行的方法,并且为这些品种的真伪鉴定和保护提供科学依据。

EST （Expressed Sequence Tag） is a valuable resource for developing SSR marker.In this study,1114 EST-SSR from 989 EST in NCBI （National Center for Biotechnology Information）database,representing 8.63％ of the total number of EST were identified.Among these sequences,tri-nucleotide repeat and hexa-nucleotide repeat were the most common types,and accounting for 39.86％ and 34.56％ of the total EST-SSR respectively.47 Tri-nucleotide repeat motifs and 113 Hexa-nucleotide repeat motifs were screened.49 SSR primers were designed to sequence flanking SSR,and 40 primer pairs showed the amplification,accounting for 81.63％ of total primers.Then the primers showing amplification were subjected to PCR for DNA from 24 cannabis varieties,and 29 primer pairs showed polymorphisms,accounting for 72.5％ of primers available.The fingerprint of 24 cannabis varieties was constructed by 7 pairs of primers （E-7、E-8、E-11、E-18、E-21、E-39 and E-48）.Results showed that it is an effective and feasible approach to develop SSR markers based on EST in Cannabis.Additionally,the fingerprint will provide reliable and scientific evidences for identifying variety and protecting intellectual property rights.