神经肽Y基因在COS－7细胞中的表达

THE EXPRESSION OF NPY cDNA IN COS-7 CELLS

构建大鼠ＮＰＹ基因的瞬时表达载体ｐＳＶＬ－ＮＰＹ，以Ｌｉｐｏｆｅｃｔｉｎ^（ＴＭ）介导将其导入ＣＯＳ－７细胞中，Ｎｏｒｔｈｅｒｎｂｌｏｔ和ＨＰＬＣ检测结果显示：ｐＳＶＬ－ＮＰＹ质粒转染的ＣＯＳ－７细胞可表达ＮＰＹｍＲＮＡ及成熟肽ＮＰＹ。

NPY cDNA fragment digested from TA-NPY was subcloned into the transient expression vector pSVL to construct the recombinant plasmid pSVL-NPY. The resultant plasmid was verified by restriction endonuclease analysis. COS-7 cell were grown to log phase and transfection were performed using Lipofectin￣(TM) reagent.Collect the supertant and cell for further analysis. Northern blot analysis indicated that NPY mRNA was exprssed in pSVLNPY transfected cells,but not in pSVL transfect cells. The result of HPLC assays of transfectant supernant show pSVL-NPY transfected cells can express NPY and the estimated level is 10.75 ng/m1/10￣7 cells.This indicated that the construction pSVL-NPY can direct the correct NPY systhesis in COS-7 cells.