WISP基因家族的克隆、表达及系统进化树的构建
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中国水产科学研究院 水产生物应用基因组研究中心,北京;上海海洋大学 水产与生命学院,上海,中国水产科学研究院 水产生物应用基因组研究中心,北京;大连海洋大学 生命科学与技术学院,辽宁 大连,中国水产科学研究院 水产生物应用基因组研究中心,北京,中国水产科学研究院 水产生物应用基因组研究中心,北京

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国家“八六三”高技术研究发展计划(2011AA100401);公益性行业(农业)科研专项项目(200903045)


Molecular cloning,expression and phylogenetic tree′s construction of WISP gene family in common carp
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The Centre for Applied Aquatic Genomics,Chinese Academy of Fishery Sciences;College of Fishery and Life Sciences, Shanghai Ocean University, Shanghai,The Centre for Applied Aquatic Genomics,Chinese Academy of Fishery Sciences;College of Aqua-Life Science and Technology, Dalian Fisheries University, Dalian,Liaoning,The Centre for Applied Aquatic Genomics,Chinese Academy of Fishery Sciences,The Centre for Applied Aquatic Genomics,Chinese Academy of Fishery Sciences

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    摘要:

    Wnt1诱导分泌蛋白(WISP)基因家族与CYR61、CTGF、NOV基因共同构成了CCN家族。研究通过鲤基因组序列与斑马鱼WISP基因编码区全序列的比对获得WISP1a、WISP1b、WISP2、和WISP3等4条序列。经过克隆、测序、比对拼接得到其开放阅读框,分别是1 089、1 077、1 038和1 026 bp,它们都是由5个外显子和4个内含子构成,分别编码362、358、345和341个氨基酸。分子系统学分析表明,鲤4个WISP基因具有高度同源性,其中WISP1a、WISP1b和WISP2均含有4个模块,WISP3缺少第2个模块。通过RT-PCR检测WISP基因在鲤组织中的表达,结果表明,WISP1a鲤在13个组织中均有表达,皮肤中表达最高,脾、肠、卵巢次之,其他组织中表达较低。WISP1b在精巢、脑、皮肤、卵巢中表达较高。WISP2在血液、鳃中表达较高。WISP3在血液、脑、肝中表达较高。实时荧光定量PCR法分析WISP基因在鲤胚胎发育时期的表达,结果表明,除WISP3外,WISP1a、WISP1b和WISP2在前期表达较高,36 h表达最低,随后逐渐升高至第6天开始下降。

    Abstract:

    Wnt1 inducible signaling pathway proteins(WISP()constitute the CCN family with CYR61,CTGF and NOV.We used BLAST program at the NCBI to identify near matches in the common carp( Cyprinus carpio)genome database to each zebrafish( Danio rerio) WISP . WISP 1a, WISP 1b, WISP 2 and WISP 3 were identified respectively.ORF of the four isforms of WISP were cloned and sequenced in common carp. WISP 1a was 1 089 bp encoding polypeptides of 362 amino acids with a calculated Mw of 41.2 ku and P I of 8.66. WISP 1b was 1 077 bp encoding a polypeptide of 358 amino acids with a calculated Mw of 41.0 ku and PI of 8.42. WISP 2 was 1 038 bp encoding polypeptides of 345 amino acids with a calculated Mw of 38.4 ku and PI of 8.22. WISP 3 was 1 026 bp encoding polypeptides of 341 amino acids with a calculated Mw of 37.9 ku and PI of 9.10.They all have 5 exons and 4 introns.Phylogenetic relationship analysis indicated that WISP 1, WISP 2 and WISP 3 were highly homologous and were clustered to their corresponding subgroup respectively. WISP 1a, WISP 1b and WISP 2 all contained 4 conservative domains and WISP 3 lost the second domain in common carp.RT-PCR analysis revealed WISP 1a to be expressed in most common carp tissues,with the principal expression in skin and weakest expression in blood cells. WISP 1b was predominantly expressed in testicle,brain,skin and ovary. WISP 2 was highly observed in blood cells and gill. WISP 3 was most strongly expressed in blood cells and also expressed in brain,liver,spleen,head kidney and not detected in other tissues.Quantitative real-time PCR were used to analyse the expression of WISP in common carp development,and the result indicated that WISP transcriptions are expressed highly at the initial stage except WISP 3 and all lowest at 36 hpf and then increased before 6th d.

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孙婷,刘伟,徐鹏,孙效文.鲤WISP基因家族的克隆、表达及系统进化树的构建[J].水产学报,2012,36(2):180~190

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  • 收稿日期:2011-10-02
  • 最后修改日期:2011-11-08
  • 录用日期:2012-02-14
  • 在线发布日期: 2012-02-16
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