人CyclinD1在毕赤酵母中的表达

Expression of Human CyclinD1 in Pichia Pastoris

目的：克隆人肝癌组织中的Cyclin D1基因，并在毕赤酵母中表达 Cyclin D1蛋白。方法从人肝癌组织中提取RNA，逆转录PCR扩增获得Cyclin D1 cDNA，将PCR产物进行TA克隆和DNA序列分析；阳性TA克隆Cyclin D1片段再亚克隆入毕赤酵母表达载体Ppic9k，甲醇诱导表达。结果逆转录 PCR扩增得到约483 bp的 DNA片段，TA克隆和DNA序列分析结果显示重组片段为人的Cyclin D1基因，Cyclin D1 cDNA亚克隆入 Ppic9k载体的 NotI和 EcoRI位点之间；甲醇诱导得到分子量约36KD的蛋白。结论成功克隆人肝癌组织中的Cyclin D1基因，并且在毕赤酵母中成功表达。

Objective To obtain the Cyclin D1,the Cyclin D1 gene was cloned and expressed in Pichia pastoris.Methods The total RNA was extracted from liver cancer tissue.The Cyclin D1 cDNA was obtained by RT-PCR.The Cyclin D1 cDNA was sequenced.And sub-clonedto the Ppic9k,the Pichia pastoris expressed was used to obtain the Cyclin D1.Results The 483 bp Cyclin D1 cDNA was obtained.And the sequence of Cyclin D1 was corrected.The 36KD Cyclin D1 was obtained by Pichia pastoris expression.Conclusion The Cyclin D1 cDNA was cloned and Cyclin D1 was expressed in Pichia pastoris.