小尾寒羊高繁殖力候选基因ESR在同期发情处理过程中的表达分析

Expression Analysis of Prolificacy-related ESRGene during Synchronized Estrus Treatment for Small Tail Han Sheep

试验选择高繁殖力小尾寒羊为试验动物,采用公认的CIDR结合外源激素处理使其同期发情,与自然发情及乏情的小尾寒羊作对比,利用反转录荧光定量RT-PCR和Western blotting技术研究雌激素受体(estrogen receptor,ESR)基因在卵巢细胞中的表达差异,同时对其启动子区的甲基化岛进行了分析。结果发现,自然发情绵羊和外源激素处理发情的羊在发情表现和时间上无明显差别;在转录水平的检测结果上也无显著性差异(P>0.05);但是它们显著高于在乏情期绵羊卵巢细胞内的表达(P<0.05)。尽管激素处理的同期发情羊卵巢细胞内的ESR蛋白表达量显著高于乏情期绵羊,但是仍然显著低于自然发情期绵羊卵巢细胞的表达(P<0.05)。此外,自然发情和激素处理的绵羊ESR基因启动子区CpG岛甲基化水平分别为0和10.77%,而发情期绵羊ESR基因启动子处于超甲基化状态(92.30%)。以上结果说明,外源激素处理的同期发情小尾寒羊尽管发情表现、时间等与自然发情的绵羊类似,但是ESR基因在蛋白水平上差异显著,且其启动子区CpG岛甲基化水平也存在显著性差异,这也许是目前胚胎移植后着床率低下的原因之一。

This study was aimed to explore the underlying reason of low implantation rate after embryo transplantation.Exogenous hormones and CIDR were used to treat Small Tail Han sheep,the estrogen receptor（ESR）gene expression in ovarian cells derived from estrus synchronization treatment（EST）,natural estrus control（NEC）and anestrus（ANE）Small Tail Han sheep were compared,and the methylation levels of CpG island was analyzed.The results showed that there was no significantly difference in estrus performance and time between EST and NET sheep,and the transcript levels of ESR were also not significant difference（P〈0.05）,but they were significantly higher than that in ANE sheep ovarian（P〈0.05）.Furthermore,though the expression amount of ESR at protein level was greatly higher in EST sheep ovarian compared to that ANE sheep（P〈0.05）,it was significantly lower than that in NEC sheep（P〈0.05）.Methylation levels at the CpG island of ESR promoter was 0and 10.77% in NEC and EST sheep,respectively;while the promoter was hypermethylation in ANE sheep（92.30%）.These results suggested that EST sheep showed similar estrus performance and time to that of NEC sheep after treated with exogenous hormones,but ESRgene expression at protein level was significantly different,and the methylation level of promoter was also significant difference.Perhaps it was one of the main causes resulting in the low implantation rate after embryo transplantation of Small Tail Han sheep.