[1]周楠楠,杨 振,罗丽芸,等.毛冠鹿AIF-1 原核载体的构建及表达[J].南京师大学报(自然科学版),2014,37(02):85.
　Zhou Nannan,Yang Zhen,Luo Liyun,et al.Prokaryotic Expression System Construction of the Tufted Deer(Elaphodus cephalophus)AIF-1[J].Journal of Nanjing Normal University(Natural Science Edition),2014,37(02):85.
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毛冠鹿AIF-1 原核载体的构建及表达()
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《南京师大学报（自然科学版）》[ISSN:1001-4616/CN:32-1239/N]

卷:
第37卷
期数:
2014年02期
页码:
85
栏目:
生命科学
出版日期:
2014-06-30

文章信息/Info

Title:
Prokaryotic Expression System Construction of the Tufted Deer(Elaphodus cephalophus)AIF-1
作者:
周楠楠; 杨 振; 罗丽芸; 宋 菲; 白 敏; 曹祥荣
南京师范大学生命科学学院,江苏省分子医学生物技术重点实验室,江苏南京210023
Author(s):
Zhou Nannan; Yang Zhen; Luo Liyun; Song Fei; Bai Min; Cao Xiangrong
School of Life Sciences,Nanjing Normal University,Jiangsu Key Laboratory for Molecular and Medical Biotechnology,Nanjing 210023,China
关键词:
毛冠鹿; AIF-鄄1; 原核表达
Keywords:
Tufted deer(Elaphodus cephalophus); AIF-1; prokaryotic expression
分类号:
Q28
文献标志码:
A
摘要:
从毛冠鹿睾丸cDNA 文库中筛选出毛冠鹿AIF-1 基因,对其进行生物信息学分析,设计引物克隆毛冠鹿AIF-1 cDNA,连入pMD19-T 载体,测序正确后酶切,与表达载体pET-28a( +) 连接,转化E. coli BL21( DE3),IPTG 诱导表达,将诱导表达重组蛋白的菌体超声破碎后,进行可溶性分析,并对可溶性蛋白进行纯化,SDS-PAGE 电泳,Western Blot 分析以及鉴定重组蛋白. 结果表明,毛冠鹿AIF-1(TdAIF-1)含有1 个438bp 的开放阅读框,编码145 个
Abstract:
The TdAIF鄄1 cDNA was cloned from the testis cDNA library of the Tufted deer(Elaphodus cephalophus) and analyzed by bioinformatic methods. Primers were designed according to cDNA sequence to clone the gene. The gene was cloned into pMD19鄄T vector for sequencing. The right sequence was digested by restriction enzyme and subcloned into the expression vector pET鄄28a( +). After transformed into E. coli BL21( DE3),the recombinant plasimid was induced to express by IPTG. The E. coli BL21(DE3)expressed recombinant protein was broken by ultrasonic to show whether the re鄄 combinant protein was soluble or not. Lastly,the soluble protein was purified. The recombinant protein was analyzed and identificated by SDS-PAGE electrophoresis and Western blot. Analysis of sequence showed that the TdAIF-1 cDNA contained a 438 bp op-n reading frame encoding 145 amino acids. The recombinant plasimid was correctly constructed according to sequencing and restriction enzyme analysis. The recombinant protein was about 20 kD and soluble mainly. When the recombinant protein was purified,using elution buffer containing 50 mmol/ L or 100 mmol/ L imidazole could get purified protein. The TdAIF-1 Prokaryotic Expression System was constructed successfully and recombinant protein was obtained,which was helpful for the future study of its biological function.

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相似文献/References:

[1]曹祥荣,王莹,李淑峰,等.毛冠鹿p16~(INK4)基因第二外显子序列分析[J].南京师大学报(自然科学版),2001,24(03):80.
　Cao Xiangrong,Wang Ying,Li Shufeng,et al.Analysis of p16INK4 Gene Exon 2 in Elaphodus Cephalophus[J].Journal of Nanjing Normal University(Natural Science Edition),2001,24(02):80.

备注/Memo

备注/Memo:
收稿日期:2013-11-25.
基金项目:国家自然科学基金(30771172)、国家基础科学人才培养基金(J1103507、J1210025).
通讯联系人:曹祥荣,教授,博导,研究方向:肿瘤发生与基因治疗分子机制. E-mail:caoxiangrong@ njnu. edu. Cn
更新日期/Last Update: 2014-06-30