大鼠IL-10真核表达载体在软骨细胞中的表达

Construction of rat IL-10 eukaryotic expression vector and its expression in rat chondrocyte

目的 构建大鼠IL-10真核表达载体并在大鼠软骨细胞中进行表达。方法 将目的基因片段通过酶切连入真核表达载体pcDNA3,并以Super Fect Transfection Reagent介导转入大鼠软骨细胞表达,RT-PCR检测细胞中mRNA水平,及ELISA检测细胞培养上清液中的IL-10蛋白含量。结果 成功构建了大鼠IL-10真核表达载体,转入软骨细胞后,检测到细胞内IL-10mRNA转录,细胞培养24、48和72 h后,经ELISA检测上清液中IL-10蛋白含量分别为36、62和100 pg/mL。结论 大鼠IL-10真核表达载体的构建并在软骨细胞中表达,为研究IL-10诱导软骨细胞同种异体移植耐受奠定了基础。

Objective To construct rat IL-10 eukaryotic expression vector and observe its expression in rat chondrocyte. Methods Target gene and vector were ligated by T4 ligase. The recombinant vectors were transfected into rat chondrocyte by Super Fect Transfection Rea-gent. IL-10 mRNA were detected by RT-PCR, and the cell supematant were assayed for rat IL-10 by ELISA method. Results Rat IL-10 eukaryotic expression vector had been constructed successfully, and had been transfected into rat chondrocyte. IL-10 mRNA was detected by RT-PCR, and cell culture supernatant was tested for IL-10 protein by ELISA in 24, 48 and 72 hours, and the results are 36, 62 and 100 pg/mL respectively. Conclusion the construction of rat IL-10 eukaryotic expression vector and its successful expression in rat chondrocyte provide a foundation for induction of immune-tolerance of allogenic chondrocyte transplantation.