茶树EST-SSRs分布特征及引物开发

Characterization of EST-derived Microsatellites and Development of SSR-markers in Tea(Camellia sinensis)

为了在茶树中开发EST-SSRs功能性标记，利用生物信息学方法对NCBI网上公开的3288奈茶树（Camellia subebsus）ESTs序列进行EST-SSRs特征分析。剔除冗余序列，得到非冗余序列2083条。在非冗余序列中发现含不同重复基元SSRs的EST序列有385条，共486个EST-SSRs，平均相隔2．10kb出现1个SSR。在2～6bp的重复基元中，二核苷酸重复基元的SSRs出现频率最高（51．97％），其次是三核苷酸（19．55％）。对所有的重复基元类型进行统计分析发现，所占比例最高的是AG／CT（47．74％），其次分别是AT／TA（4．73％）和AAG／CTT（4．73％）。利用Prime5软件，设计了206对EST-SSRs引物，随机选用72对引物进行SSR扩增，发现31对引物可以扩增出条带，其中29对引物具有多态性，多态性比率为93．5％。这些EST-SSRs将有助于茶树基因组学方面的研究。

To develop EST-SSRs functional markers in tea （Camellia sinensis） , characterization of simple sequence repeats （SSRs）were analyzed by 3288 EST （expressed sequence tag）sequences in Camellia sinensis from NCBI. A total of 2083 non-redundant EST sequences were obtained from 3288 sequences by redundant analysis and there were 486 EST-SSRs present in 385 ESTs out of the non-redundant sequences with one EST-SSR in every 2. 10 kb of EST sequence in average. Among the different motifs ranging from 2bp to 6bp,dinucleotide repeats were most abundant（51.97% ）, followed by trinucleotide repeats （ 19.55% ） and mononucleotide repeats （23.35%）. Among all identified motif types,AT/TA had the highest frequency（4. 73% ） ,followed by AAG/CTT（4. 73 % ）. 206 pairs of SSR primers were designed by the software,primer 5. Among 72 pairs of primers selected randomly from them,31 primer pairs were successfully amplifying DNA fragments with 29 polymorphic primers. The proportion of primers that produced polymorphic fragments was 93.5%. Furthermore,the availability of microsatellite markers can be expected to enhance the power and resolution of genomic analysis in tea.