[1]李文良,毛立,杨蕾蕾,等.稳定表达猪Viperin 的 PK-15 细胞系的构建与鉴定[J].江苏农业学报,2016,(01):128-132.[doi:10.3969/j.issn.1000-4440.2016.01.020]
 LI Wen-liang,MAO Li,YANG Lei-lei,et al.Construction and identification of PK-15 cell line stably expressing porcine Viperin[J].,2016,(01):128-132.[doi:10.3969/j.issn.1000-4440.2016.01.020]
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稳定表达猪Viperin 的 PK-15 细胞系的构建与鉴定()
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江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2016年01期
页码:
128-132
栏目:
畜牧兽医·水产养殖
出版日期:
2016-01-08

文章信息/Info

Title:
Construction and identification of PK-15 cell line stably expressing porcine Viperin
作者:
李文良毛立杨蕾蕾郝飞张纹纹江杰元
(江苏省农业科学院兽医研究所,农业部兽用生物制品工程技术重点实验室,国家兽用生物制品工程技术研究中心,江苏南京210014)
Author(s):
LI Wen-liangMAO LiYANG Lei-leiHAO FeiZHANG Wen-wenJIANG Jie-yuan
(Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences/Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture/National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China)
关键词:
ViperinPK-15细胞系表达
Keywords:
swineViperinPK-15 cell lineexpression
分类号:
S858.28
DOI:
10.3969/j.issn.1000-4440.2016.01.020
文献标志码:
A
摘要:
为了探讨先天免疫效应因子Viperin的抗病毒作用,从猪外周血单核细胞中成功扩增了猪Viperin基因完整阅读框,克隆到真核表达载体pEGFP-C1中构建重组表达质粒pEGFP-Vi。阳性质粒经PCR、酶切和测序鉴定后通过脂质体Lipofectamine 2000介导转染PK-15细胞,荧光显微镜观察发现融合蛋白质EGFP-Viperin定位于细胞质中,不同于对照质粒EGFP在细胞中的均匀分布。共聚焦检测结果表明,重组蛋白质主要定位于细胞的内质网上。Western blot检测结果表明,猪Viperin蛋白质在PK-15细胞中以融合蛋白质的形式稳定表达,分子量约为 7.0×104。
Abstract:
In order to explore the antiviral function of innate immunity effector Viperin, the Viperin coding region amplified from porcine peripheral blood mono nuclear cells was cloned in to expression vector pEGFP-C1. The positive plasmid, pEGFP-Vi, was detected by PCR, enzyme digestion and sequencing. PK-15 cells were transfected with pEGFP-Vi via Lipofectamine 2000. As observed by fluorescence microscope, the fusion protein EGFP-Viperin was localized in the cytoplasm, different from the uniform distribution of EGFP in the cell. Confocal laser scaning microscope rerealed that EGFP-Viperin was localized in endoplasmic reticulum. Western blot analysis showed that EGFP-Viperin fusion protein was successfully expressed with molecular weight of 7.0×104.

参考文献/References:

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备注/Memo

备注/Memo:
收稿日期:2015-07-03基金项目:江苏省自然科学基金项目(BK20130729);国家自然科学基金项目(31402180)作者简介:李文良(1984-),男,河南开封人,博士,副研究员,主要从事分子病毒学、动物疫病防治和诊断技术研究。(E-mail)kfliwenliang@163.com通讯作者:江杰元,(Tel)025-84391135;(E-mail)1776556843@qq.com
更新日期/Last Update: 2017-05-08