猪β<sub>2</sub>肾上腺素受体基因克隆及重组酵母表达质粒的构建

王健; 李永飞; 佘永新; 金茂俊; 王静

doi:10.13386/j.issn1002-0306.2015.18.023

猪β₂肾上腺素受体基因克隆及重组酵母表达质粒的构建

1. 中国农业科学院农业质量标准与检测技术研究所农业部农产品质量安全重点实验室
2. 河北北方学院食品科学系

基金项目:

国家公益性行业(农业)科研专项(201203094)；

详细信息

作者简介:
王健(1980-),男,博士研究生,副教授,研究方向:农产品质量安全快速检测技术,E-mail:xuanyuanjian0228@126.com。;

王静(1963-),女,博士,教授,研究方向:食品安全与检测技术,E-mail:w_jing2001@126.com。;

中图分类号: Q78
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出版历程
- 收稿日期: 2015-02-04

Cloning of β2 adrenergic receptors c DNA and construction of yeast expression plasmid

Institute of Quality Standards and Testing Technology for Agri-products of CAAS,Key Laboratory of Agrifood Safety and Quality,Ministry of Agriculture
Department of Food Science,Hebei North University

摘要

摘要: 克隆猪β2AR基因并进行序列分析,构建其重组酵母表达质粒。采集新鲜猪肝组织,提取总RNA,经RT-PCR扩增、克隆及鉴定,获得猪β2AR的c DNA序列,并对该序列及编码的氨基酸序列进行分析。同时将目的基因插入p PICZαA酵母表达载体,构建重组表达质粒。克隆获得猪β2AR基因c DNA序列1257 bp,Genbank登录号为KF023571.1,编码418个氨基酸,蛋白分子质量46.73 ku。生物信息学分析表明,该基因与其他物种的β2AR基因相似性较高,均在83%以上,具有较高的同源性。经PCR和双酶切鉴定以及测序分析,证明成功构建了重组酵母表达质粒p PICZαA-β2AR。本研究为猪β2AR基因在毕赤酵母系统的表达及建立基于受体的β激动剂多残留检测方法奠定了基础。
- β2肾上腺素受体 /
- 克隆 /
- 酵母表达质粒 /
- 猪
Abstract: This study aimed to clone pig β2AR gene,analysize its sequence and construct its recombinant yeast expression plasmid. Total RNA was extracted from pig liver, and then RT- PCR was conducted based on published pig β2AR gene sequence(AF000134). By cloning of the PCR product and identification,the c DNA fragment of pig β2AR was obtained. Bioinformatical tools were adopted to analyze the gene sequence and amino acid. The target gene was cloned to p PICZα A vector,constructing recombinant expression plasmid.The c DNA fragment of pig β2AR(KF023571.1) was 1257 bp,and encodeed 418 amino acids. The deduced protein was predicted to have a computed molecular mass of 46.73 ku. There were high similarity(more than83%) and homology in β2AR gene between pig and other species. Finally,recombinant plasmid named p PICZαA- β2AR was constructed successfully after verification by PCR, restriction enzyme analysis and sequencing. This research will serve as a base for expression of pig β2AR gene in yeast expression system and its application in multiresidue determination of β2-adrenoceptor agonists.
- β2adrenergic receptor /
- clone /
- yeast expression plasmid /
- pig

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