文蛤肠、外套膜和肝胰脏组织cDNA文库构建及其ESTs序列分析  

Construction of cDNA libraries from intestine,mantle and hepatopancreas of hard clam (Meretrix meretrix) and ESTs analysis

作　　者：高祥刚[1] 李云峰[1] 宋文涛[1,2] 王健[1,2] 傅立元[1,2] 刘卫东[1] 鲍相渤[1] 赫崇波[1] 

机构地区：[1]辽宁省海洋水产科学研究院,辽宁省海洋水产分子生物学重点实验室,辽宁大连116023 [2]辽宁师范大学生命科学学院,辽宁大连116029

出　　处：《中国水产科学》2010年第2期344-350,共7页Journal of Fishery Sciences of China

基　　金：辽宁省科学技术计划重大、重点项目(2008203001);辽宁省海洋与渔业科研计划项目(200801);国家海洋公益性行业科技专项(200805037)

摘　　要：利用SMART cDNA文库构建试剂盒构建了文蛤(Meretrix meretrix)肠、外套膜和肝胰脏组织的cDNA文库。经测定原始文库滴度分别为2.10×106、1.70×106和1.60×106,重组率高于95%,肠组织文库插入片段长度均大于1000bp,外套膜和肝胰脏组织文库的插入片段长度大于1kb的占87.5%,文库质量符合标准要求。随机选取文库的3168个克隆进行5′端测序,获得高质量表达序列标签(Expressed Sequence Tags,ESTs)3029个(肠:1005,外套膜:1019,肝胰脏:1005),测序成功率95.58%。经质量控制和拼接得到1796个单基因簇(Unigene),其中306个叠联群(Contigs),1490个单一序列(Singletons)。通过Blastx搜索比对、查询和注释分析,共得到已知基因696个(肠:216,外套膜:235个;肝胰脏:245个),占总数38.75%。在1796个单基因簇(Unigene)发现微卫星序列55条,这些存在微卫星位点的序列占整个ESTs数据库的3.1%。The cDNA libraries of intestine, mantle and hepatopancreas of hard clam （Meretrix meretrix）were constructed with SMART cDNA Library Construction Kit. The libraries have a high titer of 2.10 × 10^6, 1.70 × 10^6 and 1.60 × 10^6, respectively. The recombined efficiency of each cDNA library exceeded 95 %. The all insert sizes of intestine library were over 1 kb, and the insert sizes over 1 kb in the mantle and hepatopancreas libraries were over 87.5%. Atotal of 3 168 clones of the libraries were randomly picked and sequenced from the 5 ＇ end of the cDNAs using a M13 universal primer, and 3 029 raw sequences of the ESTs were processed and then assembled into 1 796 unigene involving 306 contigs and 1 490 singletons. Blastx analysis showed that 696 unigenes （ 216 for intestine, 235 for mantle and 245 for hepatopancreas） （ 38.75% ） had significant homology （ Evalue ≤ 10^-5 ） to genes with known or putative functions in GenBank.The sequences containing microsatellite DNA （SSR） accounted for 3.1% of all ESTs indicating that SSR sequences were rich in the ESTs of hard clam. [Journal of Fishery Sciences of China, 2010,17（ 2 ） ： 344-350]

关 键 词：文蛤 CDNA文库 表达序列标签 微卫星标记 

分 类 号：S959[农业科学—水产养殖]