真核表达载体pIRES2-AcGFP1-PLIN2的构建及其在3T3-L1前脂肪细胞中的表达

doi:10.19640/j.cnki.jtau.2018.02.014

天津农学院学报 ›› 2018, Vol. 25 ›› Issue (2): 55-59.doi: 10.19640/j.cnki.jtau.2018.02.014

真核表达载体pIRES2-AcGFP1-PLIN2的构建及其在3T3-L1前脂肪细胞中的表达

赵小琪, 黄英, 杨明华, 潘洪彬, 赵素梅^通信作者

云南农业大学动物营养与饲料重点实验室,昆明 650201

收稿日期:2018-03-26 出版日期:2018-06-20 发布日期:2019-11-12
通讯作者: 赵素梅（1974–）,女,教授,博士,主要从事动物分子营养与代谢调控方面的研究。E-mail：zhaosm2009@126.com。
作者简介:赵小琪（1993–）,男,硕士在读,主要从事动物分子营养与代谢调控方面的研究。E-mail：583478872@qq.com。
基金资助:
国家自然科学基金项目（31760645,31260592,31060331）

Construction of eukaryotic expression plasmid pIRES2-AcGFP1- PLIN2 and its expression in 3T3-L1 preadipocytes

ZHAO Xiao-qi, HUANG Ying, YANG Ming-hua, PAN Hong-bin, ZHAO Su-mei^{Corresponding Author}

Key Lab of Agricultural Animal Nutrition and Feed Science, Yunnan Agricultural University, Kunming 650201, China

Received:2018-03-26 Online:2018-06-20 Published:2019-11-12

摘要/Abstract

摘要： 构建pIRES2-AcGFP1-PLIN2真核表达载体,并观察其在3T3-L1前脂肪细胞中的转染状况。采用分子克隆技术,将PLIN2基因连接到pMD18-T载体中做EcoRⅠ/Sal I双酶切,将酶切产物回收并将双酶切后的目的片段与pIRES2-AcGFP1载体连接,转入DH5α感受态细胞,并提取pIRES2-AcGFP1-PLIN2阳性重组质粒,转染到3T3-L1前脂肪细胞,显微镜观察转染情况。本研究成功克隆出了PLIN2基因,同时成功构建了pIRES2-AcGFP1-PLIN2真核表达载体,并在3T3-L1前脂肪细胞中成功表达,于显微镜下观察到绿色荧光,试验为进一步研究PLIN2基因对脂肪细胞的调控机理奠定了基础。

关键词: 乌金猪, PLIN2基因, 真核表达载体

Abstract: The eukaryotic expression plasmid pIRES2-AcGFP1-PLIN2 was constructed and its transfection status in 3T3-L1 preadipocytes was observed. The PLIN2 gene was cloned into pMD18-T vector and digested by EcoR I/Sal I. The digested product was digested and then ligated into pIRES2-AcGFP1 vector and transformed into DH5α competent cells, and pIRES2-AcGFP1-PLIN2 positive recombinant plasmid was extracted and transfected into 3T3-L1 preadipocytes. The Transfection condition was observed by microscope. The PLIN2 gene was successfully cloned. At the same time, the eukaryotic expression plasmid pIRES2-AcGFP1-PLIN2 was successfully constructed and expressed in 3T3-L1 preadipocytes. The green fluorescence was observed under the microscope, which laid the foundation for further studying the regulation mechanism of PLIN2 gene on adipocytes.

Key words: Wujin pig, PLIN2 gene, eukaryotic expression plasmid

中图分类号:

S811.3

赵小琪, 黄英, 杨明华, 潘洪彬, 赵素梅. 真核表达载体pIRES2-AcGFP1-PLIN2的构建及其在3T3-L1前脂肪细胞中的表达[J]. 天津农学院学报, 2018, 25(2): 55-59.

ZHAO Xiao-qi, HUANG Ying, YANG Ming-hua, PAN Hong-bin, ZHAO Su-mei. Construction of eukaryotic expression plasmid pIRES2-AcGFP1- PLIN2 and its expression in 3T3-L1 preadipocytes[J]. Journal of Tianjin Agricultural University, 2018, 25(2): 55-59.

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http://xuebao.tjau.edu.cn/CN/Y2018/V25/I2/55

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真核表达载体pIRES2-AcGFP1-PLIN2的构建及其在3T3-L1前脂肪细胞中的表达

Construction of eukaryotic expression plasmid pIRES2-AcGFP1- PLIN2 and its expression in 3T3-L1 preadipocytes

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