脑心肌炎病毒内部核蛋白体进入位点介导白介素-12基因在B16F1细胞中的表达

Expression of Subunits,p40 and p35, of Interleukin 12 Mediated by Internal Ribosome Entry Site of Encephalomyocarditis Virus in B16F1 Cells

目的 研究脑心肌炎病毒的内部核蛋白体进入位点连接的鼠白介素 12 (mIL 12 )的 p4 0和p35双亚基基因在同一启动子的调控下能否得以有效地表达并形成异二聚体 p70。 方法 采用脂质体法将表达经脑心肌炎病毒的内部核蛋白体进入位点连接的p4 0和 p35的质粒mIL - 12 /AD1 1转染B16F1细胞 ,连续 3d收集并换细胞培养液 ,并检测其中的mIL 12 (p70 )量。结果 ELISA结果显示 ,mIL12 (p70 )的表达量随着时间推移逐步下降 ,由第 1天的 15 .5ng/ml逐步下降到第 3天的 9.3ng/ml。表明mIL 12 p4 0和p35双亚基基因得到有效的表达并加工形成了异二聚体p70。结论 脑心肌炎病毒的内部核蛋白体进入位点连接的双基因p4 0和 p35在同一启动子的调控下得以同时表达。

Objective To explore if cDNAs of two subunits, p40 and p35 of interleukin 12 ligated by internal ribosome entry site(IRES)of encephalomyocarditis virus(EMCV)can be expressed efficiently under the control of one promoter and processed into heterodimer p70.Methods Cationic lipid was used to deliver the plasmid, mIL 12/AD1 1,which expressed p40 and p35 ligated by EMCV IRES,into B16F1 cells. The culture medium of B16F1 cells was collected and replaced each day for 3 days and mIL 12(p70) expression level was assayed with ELISA method.Results mIL 12(p70)was efficiently expressed in B16F1 cells and the expression level declined from 15.5ng/ml on day 1 to 9.3ng/ml on day 3.Conclusion Two cDNAs of p40 and p35 ligated by EMCV IRES were expressed efficiently simultaneously under the control of one promoter.The translation mechanism of the genes between the upstream and downstream of EMCV IRES was different.