目的:寻找可能与锌指转录因子ZNF580存在相互作用的蛋白.方法:以ZNF580基因开放阅读框作为模版,PCR扩增后连接入酵母表达质粒pGB.诱饵质粒pGB-ZNF580经测序验证后转化酵母菌株Y190,人胎脑cDNA文库亦转化到能稳定表达诱饵蛋白的Y190酵母菌株中,并铺到含有营养缺陷型培养基(SD/-Trp/-His/-Leu)的培养皿上进行初步筛选.所得阳性克隆再进行β-半乳糖苷酶克隆转移滤纸实验进一步去除假阳性.随机挑取部分阳性克隆,逐一转化入含有诱饵质粒的Y190 酵母菌进行一对一验证.分离阳性克隆质粒测序,并应用生物信息学方法分析阳性克隆cDNA编码的蛋白.结果:确定了14种与ZNF580可能存在相互作用的蛋白.结论:初步探讨了ZNF580的功能及其可能参与的信号转导通路,为进一步研究转录因子ZNF580参与动脉粥样硬化的机制奠定了实验基础.%AIM; To idenlify the possible proleins inleracling with zinc finger Iranscriplion faclor ZNF580. METHODS: The open reading frame of ZNF580 gene was amplified by PCR and ligaled inlo plasmid pGB. The bail plas-mid pGB - ZNF580 was transformed inlo Y190 yeasl slrain afler sequencing. Human felal brain cDNA library was also transformed inlo the bail yeasl Y190 that could express the bail prolein. The co - Iransformanls were plaled onlo SD/ - Trp/ - Leu/ - His selective medium. Recognition of positive colonies was performed by (3 - galaclosidase colony - lifl filler assay. Some posilive plasmids were Lransformed inlo the bail yeasl separately furlher lo eliminate the false positive ones. The inserted cDNAs were exlracled and analyzed for homology using the BLAST program againsl the GenBank/EMBL/DDBJ non - redun-danl sequence database. RESULTS: Fourteen proteins were idenlified lo possibly interact with ZNF580. CONCLUSION: The possible signal palhway in which ZNF580 may be involved was initially explored.
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