p27^(KIP1)-绿色荧光蛋白融合基因在HCC-9204细胞中的瞬时表达  

The expression and localization of p27^(KIP1)-GFP fusion gene in hepatocellular carcinoma cell lines

作　　者：郑建勇[1] 李江[2] 李开宗[3] 王文亮[2] 王为忠[1] 

机构地区：[1]第四军医大学西京医院普二科,陕西西安710032 [2]第四军医大学基础部病理学教研室,陕西西安710032 [3]第四军医大学西京医院普一科,陕西西安710032

出　　处：《细胞与分子免疫学杂志》2003年第3期218-220,共3页Chinese Journal of Cellular and Molecular Immunology

摘　　要：目的 :观察 p2 7KIP1 绿色荧光蛋白 (GFP)融合蛋白在肝癌细胞系HCC 92 0 4中的表达及定位。方法 :根据p2 7KIP1基因的编码顺序 ,应用PCR技术将 p2 7KIP1基因 3′末端终止密码TAA突变为TGT ,并通过基因重组技术将其与GFP基因融合。再采用脂质体转染法 ,将 p2 7KIP1 GFP融合基因转入HCC 92 0 4细胞中 ,用荧光显微镜观察其表达情况。结果 :DNA序列测定的结果显示 ,p2 7KIP1基因与基因库中已收录的 p2 7KIP1的序列相比较 ,仅有 1个核苷酸的差异。荧光显微镜显示 ,在转染GFP基因的HCC 92 0 4 /GFP细胞中 ,荧光均匀地分布于整个细胞 ;而在转染融合基因 p2 7KIP1 GFP的HCC 92 0 4 /GFP p2 7细胞中 ,绿色荧光主要见于核内。结论 :转染的HCC 92 0 4细胞能高效表达融合基因p2 7KIP1 GFP ,且定位于细胞核内 ,与 p2 7KIP1基因的表达方式相同。AIM: To study the expression and localization of p27 KIP1 GFP fusion protein in hepatocellular carcinoma cell line HCC 9204. METHODS: According to the encoded sequence of human p27 KIP1 gene, stop codon TAA located at 3′ terminus of p27 KIP1 gene was mutated into TGT by PCR and fused with GFP gene. Then the p27 KIP1 GFP cDNA was transfected into HCC 9204 cells. The expression of p27 KIP1 GFP fusion protein was observed under fluorecence microscope. RESULTS: DNA sequencing demonstrated that difference between p27 KIP1 gene and that in genebank was only one nucleoside and the encoded protein sequence is identical. The observation under fluorecence microscope reporced showed that fluorochrome was spread in entire HCC 9204 cells transfected with GFP gene, whereas green fluorescence was seen mainly in the nucleus of HCC 9204 cells transfected with p27 KIP1 GFP gene. CONCLUSION: The human p27 KIP1 GFP fusion protein could be expressed in HCC 9204 cells and located in the nucleus, which is similar to the expression of p27 KIP1 gene.

关 键 词：肝癌细胞 P27^KIP1基因 绿色荧光蛋白 

分 类 号：Q786[生物学—分子生物学]