猪JHDM2A基因的克隆及表达分析

Cloning and Expression Analysis of Porcine JHDM2A Gene

试验旨在克隆猪JHDM2A基因,并研究其在猪卵巢组织中的表达情况。首先克隆猪JHDM2A基因,并构建pLVX-IRES-ZsGreen1-JHDM2A真核表达载体,同时对JHDM2A基因在猪卵泡发育过程中的表达情况进行分析。结果显示,克隆得到的猪JHDM2A基因编码区长度为3 945bp,编码1 315个氨基酸。通过多重氨基酸序列比对发现,猪JHDM2A基因与黄牛、水牛、绵羊和人相应氨基酸序列的同源性分别为93.5%、94.7%、94.7%和93.8%。蛋白质分子系统进化树分析结果表明,JHDM2A基因在物种进化过程中高度保守。通过脂质体转染法将构建的pLVX-IRES-ZsGreen1-JHDM2A真核表达载体导入HEK293T细胞,可观察到清晰的绿色荧光蛋白表达。免疫组化结果显示,JHDM2A蛋白在不同发育阶段猪卵泡中均有表达。本试验通过克隆获得猪JHDM2A基因序列,JHDM2A蛋白在猪卵巢中高度表达,表明其功能可能与猪卵泡发育密切相关。

The aim of this study was to clone the JHDM2A gene of porcine and study the expres- sion of JHDM2A gene in porcine ovary. In this study, we cloned the porcine JHDM2A gene and constructed its eukaryotic expression vector, the expression of JHDM2A gene in porcine ovarian tissue was also analyzed. The results showed that the cloned CDS length of porcine JHDM2A gene was 3 945 bp, which encoded 1 315 amino acids. The results of multiple amino acid sequence comparison showed that the porcine JHDM2A shared 93.5%, 94.7%, 94.7% and 93.8% ho- mologous compared with those of Bubalus bubalis , Bos taurus, Ovis aries and Homo sapiens, re- spectively. Phylogenetic tree analysis indicated that the JHDM2A gene was highly conservative in the evolutionary process. The pLVX-IRES-ZsGreenl-JHDM2A eukaryotic expression vector was constructed, and clear green fluorescent signal was observed when the plasmid was transfect- ed into HEK293T cells by liposome method. The immunohistochemical results showed that the JHDM2A protein was expressed in porcine follicle of different development stages. The results showed that the porcine JHDM2A gene sequences was cloned, and the JHDM2A protein was highly expressed in porcine ovary, indicating that its function might be closely related to the de- velopment of porcine follicular.