期刊论文

Cloning and Expression of Porcine ELF4Gene and Preparation of Its Polyclonal Antibody

中文

畜牧兽医学报

0366-6964

2017

48

3

425-435

10.11843/j.issn.0366-6964.2017.03.005

国家自然科学基金青年基金（31101837） 河南省重点科技攻关项目（142102110101） 衡阳师范学院***专项项目 河南省高等学校重点科研项目（16A230023）

本校为其他机构

本研究旨在克隆猪转录因子ELF4基因,体外表达ELF4蛋白并制备抗该蛋白的多克隆抗体。采用RTPCR扩增猪ELF4基因,运用生物信息学软件分析其核苷酸和编码氨基酸序列特征,将其克隆至原核表达载体 pET28 a,构建重组表达载体pET28 a-pELF4,转化Rosetta(DE3)菌株,在不同温度、IPTG浓度、诱导时间等条件下进行诱导表达,采用SDS-PAGE和Western blot分析蛋白表达,重组pELF4蛋白(rpELF4)经纯化免疫鸡制备抗 pELF4抗体。结果表明,pELF4开放阅读框为1 989 bp(GenBank No.:KU097322),编码662个氨基酸,该基因与已经测定的人、牛、小鼠的ELF4的核苷酸相似性依次为89.4%、90.7%和81.1%,与牛等大动物的亲缘关系相对...

This research aimed to clone and express the porcine transcription factor E74-1ike factor 4 (pELF4) gene in vitro and to prepare its polyclonal antibody.The pELF4 gene was cloned by RT-PCR (Reverse Transcription-Polymerase Chain Reaction),and the characteristics of its nucleotides and coding amino acids were analyzed by the bioinformatics softwares.The recombinant expression vector pET28a-pELF4 was constructed by inserting the pELF4 gene into the prokaryotic expression vector pET28a.Then pET28a-pELF4 was transformed into E.coli Rosetta(DE3) and...