ⅡTA基因短发夹RNA表达载体对MHC-Ⅱ基因表达的抑制作用
he inhibiting effect of shRNA targeting CⅡTA on MHC-Ⅱ expression
目的：探讨大鼠CⅡTA基因的shRNA质粒载体对于CⅡTA基因和MHC-Ⅱ基因表达的抑制效果。方法：构建针对大鼠CⅡTA基因的三个shRNA质粒载体，脂质体法体外转染大鼠树突状细胞(DC)，门静脉注射法体内转染大鼠脾脏，采用荧光实时定量PCR技术检测转染后DC和脾脏的CⅡTA及MHCⅡ的mRNA表达水平变化，流式细胞术检测转染后MHC-Ⅱ的蛋白表达变化。结果：成功构建CⅡTA-shRNA质粒载体；与对照组相比，三个shRNA质粒转染组的DC和脾脏转染后的CⅡTA和MHC-Ⅱ的mRNA表达水平及MHC-Ⅱ抗原表达水平均显著性减低（P＜0.01），CⅡTA与MHC-Ⅱ基因表达具有明显正相关；pCⅡTA1-shRNA干预组对目的基因表达抑制最为明显。结论：应用CⅡTA-shRNA质粒载体显著抑制CⅡTA基因表达，RNA干扰CⅡTA可明显抑制MHC-Ⅱ基因表达。
Objective: To explore the inhibiting effect of shRNA plasmid vector targeting rat CⅡTA gene on CⅡTA and MHC-Ⅱ expression. Methods: Three shRNA plasmid vectors were constructed. Rat dendritic cells (DC) were transfected in vitro and in vivo transfection of rat spleen was performed by portal vein injection. Real time QRT-PCR and flow cytometry (FCM) were used to detect the expression of CⅡTA and MHC-Ⅱ in DC and spleen after transfection. Results: CⅡTA-shRNA plasmids were successfully constructed. Compared to control groups, the transcription levels of CⅡTA and MHC-Ⅱ and the expression level of MHC-Ⅱ were significantly inhibited in all three pCⅡTA-shRNA experimental groups (P＜0.01). There was positive correlation between the expression of CⅡTA and MHC-Ⅱ. Among the three pCⅡTA-shRNA groups, pCⅡTA1-shRNA showed the strongest inhibiting effect. Conclusion: The expressions of CⅡTA and MHC-Ⅱ are inhibited after pCⅡTA-shRNA transfection. shRNA interfering CⅡTA significantly downregulates the MHC-Ⅱ expression.
何向辉、章志翔、赵娜、刘刚、朱理玮
基础医学生物科学研究方法、生物科学研究技术分子生物学
ⅡTAMHC-ⅡshRNA质粒载体
Key words: CⅡTAMHC-ⅡshRNAplasmid vector
何向辉,章志翔,赵娜,刘刚,朱理玮.ⅡTA基因短发夹RNA表达载体对MHC-Ⅱ基因表达的抑制作用[EB/OL].(2008-03-24)[2025-05-22].http://www.paper.edu.cn/releasepaper/content/200803-694.点此复制
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