绵羊肺腺瘤病毒囊膜蛋白的表达可促进NIH3T3细胞增殖  

Envelope protein of Jaagsiekte sheep retrovious expressed in NIH3T3 cells promotes cell proliferation

作　　者：杜方原 陈大勇 张宇飞[1] 孙晓林[1] 郭文庆[1] 刘淑英[1,3] 

机构地区：[1]内蒙古农业大学兽医学院,内蒙古呼和浩特010018 [2]内蒙古赛诺草原羊业有限公司,内蒙古呼和浩特010040 [3]农业部动物临床诊疗技术重点实验室,内蒙古呼和浩特010018

出　　处：《细胞与分子免疫学杂志》2016年第9期1188-1192,共5页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(31160493;31360597);内蒙古科技厅应用研究项目(201502070);内蒙古草原英才创新团队项目(20151031)

摘　　要：目的研究外源性绵羊肺腺瘤病毒囊膜蛋白(ex JSRV Env)对NIH3T3小鼠成纤维细胞增殖的影响。方法构建含完整的外源性绵羊肺腺瘤病毒囊膜基因(ex ISRV-env)的重组质粒pc DNA4/myc-His/ex JSRV-env,并鉴定其正确性。采用脂质体转染技术pc DNA4/myc-His/ex JSRV-env瞬时转染NIH3T3细胞,利用反转录PCR和Western blot法检测Env的表达,利用CCK-8法和平板克隆形成实验检测Env表达对NIH3T3细胞增殖的影响。结果成功构建了含ex JSRV-env的重组真核表达质粒pc DNA4/myc-His/ex JSRV-env;以重组质粒瞬时转染NIH3 T3细胞,检测到JSRV Env蛋白的表达,CCK-8法和平板克隆形成实验结果表明转染pc DNA4/myc-His/ex JSRV-env的细胞增殖速度显著高于对照组(空白及阴性)细胞。结论 NIH3T3细胞表达的JSRV Env蛋白可促进NIH3 T3细胞的增殖。Objective To explore the influence of the exogenous Jaagsiekte sheep retrovious （exJSRV） envelope protein （Env） on NIH3T3 cell proliferation. Methods A recombinant plasmid pcDNA4/myc-His/exJSRV-env carrying exJSRV-env gene was constructed, and then the correctness of the recombinant plasmid was identified by PCR, restriction enzyme digestion and sequencing. The recombinant plasmid pcDNA4/myc.His/exJSRV.env was transiently transfected into NIH3T3 cells by LipofectamineTM LTX. After the trensfection of the recombinant plasmid, the expression of exJSRV-env was detected by reverse transcription PCR and Western blotting. The effect of Env on cell proliferation was investigated by CCK-8 assay and plate colony formation assay. Results The recombinant eukaryotic expression plasmid containing exJSRV-env was successfully constructed as identified by PCR, restriction enzyme identification and sequencing. After the recombinant plasmid was transiently transfected into NIH3T3 cells, reverse transcription PCR and Western blotting showed the expression of exJSRV-env, and Env promoted NIH3T3 cell proliferation significantly. Conclusion JSRV Env was expressed successfully in the NIH3T3 cells and promoted the proliferation of NIH3T3 cells.

关 键 词：绵羊肺腺瘤病毒 囊膜蛋白 细胞增殖 平板克隆形成实验 

分 类 号：R392-33[医药卫生—免疫学] Q253[医药卫生—基础医学]