日本囊对虾Ran基因的克隆表达与蛋白质GTP结合活性分析

CORC > 集美大学

	日本囊对虾Ran基因的克隆表达与蛋白质GTP结合活性分析
	韩芳 ; 王志勇
刊名	http://epub.edu.cnki.net/grid2008/brief/detailj.aspx?filename=jmxz201004000&dbcode=CJFQ&dbname=CJFQ2010
	2012-06-05 ; 2012-06-05
关键词	日本囊对虾 Ran基因 RACE-PCR 蛋白表达 GTP活性 shrimp Ran gene RACE-PCR protein expression GTP-binding activity Q78
其他题名	Cloning,Recombinant Expression and GTP-binding Activity Analysis of Ran Gene of Penaeus japonicus
中文摘要	在日本囊对虾抑制性差减杂交(suppression subtractive hybrid ization,SSH)研究过程中,首次发现一段经同源比较为Ran基因的部分序列,在抗病日本囊对虾中上调表达.为了进一步探索日本囊对虾Ran基因的功能,通过RACE-PCR的方法克隆得到了日本囊对虾Ran基因全长共1 441个碱基,其中开放阅读框为645个碱基,共编码215个氨基酸,这是首次在海洋无脊椎动物体内克隆到该基因.还将该基因克隆到原核表达载体PGEX-4T-2中并转化大肠杆菌BL21,37℃下诱导6 h,超声裂解表达菌株,结果表明GST-Ran融合蛋白在大肠杆菌中为可溶性表达,蛋白大小约为50 ku,经纯化得到了纯度大于90%的GST-Ran融合蛋白.随后的GTP结合试验验证了Ran蛋白具有GTP结合活性.; In previous studies,a cDNA fragment was found to be highly homologous with ras-related nuclear proteins(Ran proteins)by suppression subtractive hybridization(SSH)using WSSV-resistant shrimp.In this investigation,Ran gene was cloned from Penaeus japonicus shrimp by RACE-PCR,the total cDNA was 1 441 base pair and the ORF of shrimp Ran gene contained 645 bp,which encoded 215 amino acids.This was the first Ran gene obtained from marine invertebrates.Then the gene was cloned in pGEX-4T-2 and expressed in Escherichia coli BL21.After induction with IPTG at 37 ℃ for 6 h,the GST-Ran fusion protein(about 50 ku)was expressed and purified by glutathione-agarose beads.Based on protein domain analysis,the Ran protein was predicted to have a GTP/ATP binding motif.To test whether the Ran protein was endowed with GTPase activity,the GTP-binding assay was performed.The result revealed that the Ran protein had the GTP-binding activity.; 【作者单位】集美大学水产学院；福建省高校水产科学技术与食品安全重点实验室；【作者英文名】HAN Fang1,2,WANG Zhi-yong1,2 (1.Fisheries College,Jimei University,Xiamen 361021,China;2.Key Laboratory of Science and Technology for Aquaculture and Food Safety(Jimei University),Fujian Province,Xiamen 361021,China)
语种	中文
内容类型	期刊论文
源URL	[http://ir.calis.edu.cn/hdl/235041/12199]
专题	集美大学
推荐引用方式 GB/T 7714	韩芳,王志勇. 日本囊对虾Ran基因的克隆表达与蛋白质GTP结合活性分析[J]. http://epub.edu.cnki.net/grid2008/brief/detailj.aspx?filename=jmxz201004000&dbcode=CJFQ&dbname=CJFQ2010,2012, 2012.
APA	韩芳,&王志勇.(2012).日本囊对虾Ran基因的克隆表达与蛋白质GTP结合活性分析.http://epub.edu.cnki.net/grid2008/brief/detailj.aspx?filename=jmxz201004000&dbcode=CJFQ&dbname=CJFQ2010.
MLA	韩芳,et al."日本囊对虾Ran基因的克隆表达与蛋白质GTP结合活性分析".http://epub.edu.cnki.net/grid2008/brief/detailj.aspx?filename=jmxz201004000&dbcode=CJFQ&dbname=CJFQ2010 (2012).