新基因p7TP3剪切体1的克隆与亚细胞定位

利用RT-PCR技术扩增丙型肝炎病毒p7蛋白反式调节基因3剪切体1（p7TP3剪切体1）开放阅读框序列。将目的基因片段插入pGEM-T载体中,经双酶切和测序鉴定后,定向克隆至荧光表达载体pEGFP-C1,EcoRI及BamHI双酶切鉴定。脂质体法转染HepG2细胞,荧光显微镜下观察确定其亚细胞定位。结果表明,p7TP3剪切体1片断长度为381 bp,与预期结果一致;亚细胞定位结果表明,该蛋白定位于细胞浆中。

Cloning and Subcellular Location of the Novel Gene p7TP3 Spliced Variant

The p7TP3 spliced variant 1 gene was amplified by RT-PCR,the PCR products was cloned into pGEM-T vector.Then it was inserted to plasmid pEGFP-C1(enhanced green fluorescent protein).To identify the subcellular location of p7TP3 spliced variant 1,the recombinant plasmid was transfected to HepG2 cells.Restricted enzymes analysis and DNA sequencing showed that the sequence of the recombinant plasmid pEGFP-C1-p7TP3 spliced variant 1 was correct,the location of p7TP3 spliced variant 1 was detected in cytoplasm.These results pave the way for study the biological function of p7TP3 spliced variant 1.