Fas和β-雌二醇受体融合基因在L929细胞中的表达及活性鉴定

Expression and functional characterization of Fas/ER fusion gene in L929 cells

利用ＰＣＲ方法将人Ｆａｓ基因的信号肽、跨膜区、胞内区与人β－雌二醇受体的激素结合结构域基因融合，并插入高效真核表达载体ｐｃＤＮＡ３。用脂质体法转染Ｌ９２９细胞，加入Ｇ４１８４周后，筛选出的转化细胞经Ｗｅｓｔ－ｅｒｎｂｌｏｔ检测证明，该融合基因在Ｌ９２９细胞中得到较高表达。ＭＴＴ法检测进一步表明，在培养基中加入β－雌二醇可有效地诱导转化细胞发生凋亡，ＩＣ５０为１０－１０ｍｏｌ／Ｌ。

The human Fas gene including signal peptide, transmembrane domain and cytoplasmic domain was fused with HBD of the human β estrogen receptor gene by PCR technique, and was then inserted into eukaryotic vector pcDNA3, L929 cells were transfected with the expression plasmid by lipofectamine mediated gene transfection. After selection with G418 for four weeks, transformants expressing the fusion protein were identified by Western blot. Apoptosis was induced in these cells by adding β estrodiol in the culture, and IC 50 was about 10 -10 mol/L on MTT assay.