转人B7.2基因的L929细胞对T细胞增殖和分泌IL-10的作用

Effect of the human B7.2 transfectant on the improvement of T cell proliferation and secretion of IL-10

应用RT-PCR法,从由LPS刺激的人的DC细胞中抽提mRNA,扩增获得编码人B7.2分子的cDNA,并将其克隆到PMD18-T载体,经PCR、酶切及测序进行鉴定确证,进而构建PEGZ-term-B7.2的真核表达载体。脂质体法共转染包装细胞293T,用含有完整病毒颗粒的293T细胞的上清感染L929细胞,经Zeocin筛选,建立稳定表达人B7.2分子的L929基因转染细胞株。并证实了B7.2基因转染细胞能有效地促进T细胞的体外增殖和促进IL-10的分泌。

The cDNA fragment encoding the hunman B7.2 was obtained from the mRNA of LPS-activated dendritic cells(DC) by using RT-PCR and cloned into the PMD18-T vector. By DNA sequencing, the cDNA was consistent with the reported human B7.2 cDNA in the GenBank, and then was inserted into the eukaryotic expression vector PEGZ-term. The recombinant plasmid was transfected into 293T cell line with Lipofectamine. The culture supernatant of 293T cell line including the complete virus particles was used to transfect L929 cell line. Selected by Zeocin after transfection, a stable cell line expressing the human B7.2 was established. It can increase the proliferation of T cells and the secretion of IL-10 efficiently.