仿刺参profilin基因全长cDNA的克隆及表达分析  

Full-length cDNA Cloning and Expression Analysis of Profilin Gene from Sea Cucumber(Apostichopus japonicus)

作　　者：董颖[1] 高杉[1] 陈仲[1] 杨爱馥[1] 姜北[1] 关晓燕[1] 王摆[1] 周遵春[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

出　　处：《中国农业科技导报》2013年第2期159-167,共9页Journal of Agricultural Science and Technology

基　　金：国家自然科学基金项目(30972272);辽宁省科技计划项目(2011203005);辽宁省博士科研启动基金(20111072)资助

摘　　要：仿刺参(Apostichopus japonicus)是我国北方沿海重要养殖品种之一。克隆了仿刺参profilin基因全长cDNA序列,并分析了基因的表达规律。该基因cDNA序列全长787 bp,5'-非翻译区(5'-untranslated region,UTR)长205 bp,3'-UTR长204 bp,开放阅读框378 bp,编码125个氨基酸,预测蛋白分子量13.4 kDa。仿刺参Profilin蛋白的PROF保守结构域中含有肌动蛋白互作位点、多聚脯氨酸结合位点和PIP2互作位点。氨基酸序列比对结果显示,仿刺参Profilin与丝盘虫(Trichoplax adhaerens)和囊舌虫(Saccoglossus kowalevskii)相似度最高,为46%。Quantitative real-time PCR结果显示,profilin mRNA在仿刺参未受精卵、受精卵、多细胞期、囊胚期、原肠期、小耳状幼体、中耳状幼体、大耳状幼体、樽型幼体、五触手幼体和稚参11个发育阶段及幼参的体壁、体腔细胞、肠道和呼吸树中均有表达;在仿刺参的不同发育阶段中,未受精卵至原肠期profilin mRNA表达量低,从小耳状幼体至稚参表达量增高;在仿刺参的不同组织中,profilin mRNA在体腔细胞中的表达量最高。经过棘皮动物免疫系统有效激活物脂多糖LPS的刺激,体腔细胞中profilin mRNA表达量显著升高,为仿刺参养殖病害的防控提供科学依据。Sea cucumber （Apostichopus japonicus） was one of the important aquaculture species in the north coast of China. The studies on function and mechanism of immune-related genes can provide a reference to the prevention of diseases in the cultured A. japonicus. In this study, a full-length eDNA sequence of profilin gene from A. japonicus was obtained by constructing eDNA libraries and sequencing analysis, which was 787 bp and includes a 205 bp 5＇- untranslated region （UTR）, 378 bp ORF encoding 125 amino acids with a conserved PROF domain, and a 204 bp 3＇-UTR; and the predicted molecular weight was 13.4 kDa. The conserved PROF domain included acting interaction sites, poly-L-proline binding sites, and PIP2-interaction sites. The deduced amino acid sequence showed 46% amino acid sequence identity to the Profilin of Trichoplax adhaerens and Saccoglossus kowalevskii. The profilin mRNA levels were detected by using quantitative real-time PCR at different developmental stages of sea cucumber, including unfertilized egg, fertilized egg, cetlulous stages, blastula, gastrula, early anricularia, auricularia, late auricularia, doliolaria, pentactu!a and juvenile. The expression patterns were also detected in different tissues of young sea cucumbers including intestines, coelomocytes, respiratory trees and body walls. The profilin expression levels were low from unfertilized eggs to gastrula, however, an increase in transcript levels was observed from early auricutaria to juvenile. The highest expression level was found in the coelomoeytes of young sea cucumbers. LPS was a potent activator of the echinoderm immune system, and after LPS challenge, the highest expression of profilin mRNA was observed in the coelomocytes.

关 键 词：仿刺参 PROFILIN 序列分析 基因表达 LPS刺激 

分 类 号：S917[农业科学—水产科学]