将增强的绿色荧光蛋白 ( enhanced green fluorescent protein,EGFP)基因插在 HCMV( hum ancytom egolovirus)启动子下游 ,构建了表达质粒 p CA13 - e G,用脂质体 L ipofectin介导分别转染 He L a细胞、Vero细胞 ,仅通过细胞传代 ,就获...将增强的绿色荧光蛋白 ( enhanced green fluorescent protein,EGFP)基因插在 HCMV( hum ancytom egolovirus)启动子下游 ,构建了表达质粒 p CA13 - e G,用脂质体 L ipofectin介导分别转染 He L a细胞、Vero细胞 ,仅通过细胞传代 ,就获得了能稳定高效表达 EGFP的绿色细胞 .比较发现 ,质粒 p CA13 - e G转染后 ,产生能高效表达 EGFP的 He L a细胞其比率高于 Vero细胞 ;EGFP高效表达对 Vero细胞的毒性大于对 He L a细胞的毒性 .本研究表明 ,绿色细胞轮廓清晰 ,由于其特有的性质 ,在用于细胞的形态观察、细胞分裂等研究时会有所作为 .展开更多
Asy (apoptosis/saibousi Yutsudo) is a novel apoptosis-inducing gene found in 1999 by Yutsudo group in Japan. In 2000, Qi Bing et al. cloned another novel gene, named hap (homo-logue of ASY protein), which encoded the ...Asy (apoptosis/saibousi Yutsudo) is a novel apoptosis-inducing gene found in 1999 by Yutsudo group in Japan. In 2000, Qi Bing et al. cloned another novel gene, named hap (homo-logue of ASY protein), which encoded the ASY interacting protein, from human lung cell line (WI-38) cDNA library by using yeast two-hybrid system. It has been proved that ASY formed homodimer in yeast and human cell line, ASY and HAP formed heterodimer in yeast cells, and both induced cell apoptosis in human tumor cell lines Sao2 and CGL4. This paper showed that HAP could form homodimer in yeast cells by yeast two-hybrid system; HAP and ASY could pro-duce heterodimer in human cell line by cross-immunoprecipitation test; by using apoptosis-testing technologies such as AnnexinV, TUNEL, DNA ladder and Flow Cytometry, the cell apoptosis in human normal or tumor cell lines transfected with hap or asy individually or cotransfected by the both was qualified or quantified. It was firstly demonstrated that ASY or HAP induced cell apop-tosis not only in human tumor cell lines, but also in human normal cell lines. Moreover, we proved that the heterodimer between ASY and HAP decreased apoptosis-inducing activity from the homodimer of ASY or HAP. It revealed that by choosing to form heterodimer or homodimer be-tween ASY and / or HAP is an important mechanism of regulating apoptosis in human cell lines.展开更多
asy gene is a novel apoptosis-inducing gene, but its mechanism is unclear. To investigate the mechanism of asy inducing apoptosis, a novel gene encoding ASY interacting protein (asyip) is isolated from human lung cell...asy gene is a novel apoptosis-inducing gene, but its mechanism is unclear. To investigate the mechanism of asy inducing apoptosis, a novel gene encoding ASY interacting protein (asyip) is isolated from human lung cell line (WI-38) cDNA library with yeast two-hybrid system. The asyip gene is constitutively expressed as two mRNA transcripts with the size of 1.8 and 2.7 kb in various human tissues at different levels. Sequence analysis of full-length cDNA reveals that the two alternative transcripts of asyip gene contain common 5’ end and different 3’ end, and share a common open reading frame encoding a polypeptide of 236 amino acids. Two protein kinase C phosphorylation sites and two casein kinase II phosphorylation sites are found in ASYIP amino acid sequence. Two highly hydrophobic regions encoding potentially two transmembrane domains are present. The ASYIP protein contains a C-terminal endoplasmic reticulum retrieval signal (Lys-Lys-Lys-Ala-Glu). Immunoprecipitation assay confirmed the interaction展开更多
文摘将增强的绿色荧光蛋白 ( enhanced green fluorescent protein,EGFP)基因插在 HCMV( hum ancytom egolovirus)启动子下游 ,构建了表达质粒 p CA13 - e G,用脂质体 L ipofectin介导分别转染 He L a细胞、Vero细胞 ,仅通过细胞传代 ,就获得了能稳定高效表达 EGFP的绿色细胞 .比较发现 ,质粒 p CA13 - e G转染后 ,产生能高效表达 EGFP的 He L a细胞其比率高于 Vero细胞 ;EGFP高效表达对 Vero细胞的毒性大于对 He L a细胞的毒性 .本研究表明 ,绿色细胞轮廓清晰 ,由于其特有的性质 ,在用于细胞的形态观察、细胞分裂等研究时会有所作为 .
文摘Asy (apoptosis/saibousi Yutsudo) is a novel apoptosis-inducing gene found in 1999 by Yutsudo group in Japan. In 2000, Qi Bing et al. cloned another novel gene, named hap (homo-logue of ASY protein), which encoded the ASY interacting protein, from human lung cell line (WI-38) cDNA library by using yeast two-hybrid system. It has been proved that ASY formed homodimer in yeast and human cell line, ASY and HAP formed heterodimer in yeast cells, and both induced cell apoptosis in human tumor cell lines Sao2 and CGL4. This paper showed that HAP could form homodimer in yeast cells by yeast two-hybrid system; HAP and ASY could pro-duce heterodimer in human cell line by cross-immunoprecipitation test; by using apoptosis-testing technologies such as AnnexinV, TUNEL, DNA ladder and Flow Cytometry, the cell apoptosis in human normal or tumor cell lines transfected with hap or asy individually or cotransfected by the both was qualified or quantified. It was firstly demonstrated that ASY or HAP induced cell apop-tosis not only in human tumor cell lines, but also in human normal cell lines. Moreover, we proved that the heterodimer between ASY and HAP decreased apoptosis-inducing activity from the homodimer of ASY or HAP. It revealed that by choosing to form heterodimer or homodimer be-tween ASY and / or HAP is an important mechanism of regulating apoptosis in human cell lines.
文摘asy gene is a novel apoptosis-inducing gene, but its mechanism is unclear. To investigate the mechanism of asy inducing apoptosis, a novel gene encoding ASY interacting protein (asyip) is isolated from human lung cell line (WI-38) cDNA library with yeast two-hybrid system. The asyip gene is constitutively expressed as two mRNA transcripts with the size of 1.8 and 2.7 kb in various human tissues at different levels. Sequence analysis of full-length cDNA reveals that the two alternative transcripts of asyip gene contain common 5’ end and different 3’ end, and share a common open reading frame encoding a polypeptide of 236 amino acids. Two protein kinase C phosphorylation sites and two casein kinase II phosphorylation sites are found in ASYIP amino acid sequence. Two highly hydrophobic regions encoding potentially two transmembrane domains are present. The ASYIP protein contains a C-terminal endoplasmic reticulum retrieval signal (Lys-Lys-Lys-Ala-Glu). Immunoprecipitation assay confirmed the interaction
