中国农业科学院农业信息研究所 中国农学会
Agricultural Information Institute of CAAS China Association of Agricultural Science Societies
En
Ch
中国农业科学院农业信息研究所 中国农学会 主办

AI检索

订阅

返回顶部
当前位置: 首页 > 文章 > 慢病毒载体介导稳定表达CD163的PAM细胞系的建立及对PRRSV感染的研究 畜牧兽医学报 2013,44 (11) 1797-1804
Position: Home > Articles > Generation of a Porcine Alveolar Macrophage Cell Line Stably Expressing CD163by Lentiviral Vector for the Production of Porcine Reproductive and Respiratory Syndrome Virus Chinese Journal of Animal and Veterinary Sciences 2013,44 (11) 1797-1804

慢病毒载体介导稳定表达CD163的PAM细胞系的建立及对PRRSV感染的研究

作  者：
王向鹏;魏蕊芳;肖书奇;周恩民
单  位：
西北农林科技大学动物医学院兽医免疫学研究所
关键词：
CD163;慢病毒载体;猪繁殖与呼吸综合征病毒;猪肺泡巨噬细胞
摘  要：
本研究旨在通过慢病毒载体系统将猪源CD163转染入永生化的猪肺泡巨噬细胞(PAM)系(CRL-2843),建立稳定表达CD163的PAM细胞系(PAM-CD163)及验证该细胞系对PRRSV的易感性。用PCR方法从pJET1.2-CD163载体上扩增CD163基因编码区,通过酶切连接构建慢病毒表达载体pTrip-CMV-CD163-IRESpur。将该质粒与慢病毒包装质粒psPAX2和pMD2.G共转染293-T细胞,包装表达CD163的慢病毒。将收获的慢病毒在Polybrene的介导下转导至永生化PAM细胞,采用嘌呤霉素筛选和细胞有限稀释法筛选出稳定表达CD163的PAM细胞系。应用得到的细胞系进行PRRSV感染试验。经过RT-PCR、间接免疫荧光试验、Western blot和流式细胞术试验证实,筛选出1株能稳定表达CD163的PAM细胞系,命名为PAM-CD163。该株细胞对PRRSV高度易感,PRRSV在PAM-CD163细胞上的毒价可以达到105.0 TCID50·mL-1以上。建立了稳定表达CD163的PAM细胞系,可以用于PRRSV的分离培养和细胞受体的研究。
译  名：
Generation of a Porcine Alveolar Macrophage Cell Line Stably Expressing CD163by Lentiviral Vector for the Production of Porcine Reproductive and Respiratory Syndrome Virus
作  者：
WANG Xiang-peng;WEI Rui-fang;XIAO Shu-qi;ZHOU En-min;Veterinary Immunology Institute,College of Veterinary Medicine,Northwest A&F University;
关键词：
CD163;;lentiviral vector;;porcine reproductive and respiratory syndrome virus;;porcine alveolar macrophage
摘  要：
The objectives of this study were to use the lentiviral vector pTrip-CMV-IRES-pur plasmid to deliver porcine CD163into a porcine alveolar macrophage(PAM)cell line(CRL-2843),to generate a cell line stably expressing CD163(designated PAM-CD163),and to evaluate its permissibility for PRRSV infection.The porcine CD163coding sequences were amplified by PCR from pJET1.2-CD163plasmid and inserted into downstream of CMVpromoter in the lentiviral vector pTrip-CMV-IRES-pur plasmid.Monolayer of 293-T cells were cotransfected with three plasmids psPAX2,pMD2.G and pTrip-CMV-CD163-IRES-pur.The recombinant lentivirus expressing CD163was harvested in the culture fluid.For transduction,the immortalized PAM cells(CRL-2843)were exposed to lentivirus in the presence of polybrene.The transduced cells were selected with puromycin and single cell colonies were isolated and expanded for PRRSV infection assay.The CD163gene was transcribed by RT-PCR and the protein was expressed as identified by indirect immunofluorescence assay,Western blot and flow cytometry analyses.A PAM cell line(CRL-2843)stably expressing CD163(designated PAM-CD163)was susceptible to PRRSV infection with the titer of 105.0 TCID50 ·mL-1.A PAM cell line stably expressing CD163was constructed and permissive to PRRSV infection.This cell line could be a valuable tool for PRRSV propagation and PRRSV cellular receptors study.

相似文章

导出引用
计量
文章访问数: 78
HTML全文浏览量: 0
PDF下载量: 0

所属期刊

推荐期刊

友情链接Friendly link
农业信息研究所Agricultural information institute 农业智能知识服务平台Agricultural intelligent knowledge service platform 农业学术服务平台Agricultural Knowledge Service System 中国知网CNKI 维普网VIP 万方WANFANG DATA
联系我们Contact us

电话Tell：010-82109652-808

邮箱E-mail：agrijournal@caas.cn

服务Service
关于我们About us 意见建议Opinions and suggest
版权说明Copyright
单位：中国农业科学院农业信息研究所 中国农学会 © 2017中国农业科学院农业信息研究所 ICP备案号：京ICP备09089781号-46