核移植胚胎甲基化过程相关调控因子表达情况研究

Study on the expression of regulatory factors in methylation of nuclear transfer

为了研究体细胞移入去核的卵母细胞后甲基化重编码程序的模式变化情况,试验对10个参与甲基化重编程的重要因子在5个发育阶段的早期核移植胚胎(配子期、S期、2-cell期、8-cell期和桑葚期)中表达量的变化模式进行了荧光定量研究。结果表明:核移植胚胎(SCNT胚胎)2-cell期之前去甲基化调节蛋白AID和APOBEC1出现低表达,引起去甲基化能力的显著下降;S期MBD1、Me CP2和MBD3三种甲基化结合蛋白出现高表达,维持了该时期基因组的高甲基化水平;MBD4基因表达下调,引起基因错配。说明甲基化相关因子的异常表达导致核移植胚胎甲基化异常,是诱发其出现发育异常或死胎的主要原因。

The aim of the present study was to analyze the changes in the pattern of the methylation reprogramming after somatic cell transfer to the denucleated oocyte. The expression level changs of 10 important factors involved in methylation reprogramming were studied in five developmental stages of early nuclear transfer embryos( gamophase,S phase,2-cell,8-cell and morula) by fluorescence quantification q PCR. The results showed that the low expression of the demethylation regulatory protein AID and APOBEC1 before the 2-cell stage of the nuclear transfer embryo( SCNT embryo) were responsible for significantly decreased demethylation ability. The over expression of MBD1,Me CP2 and MBD3 in S phase could maintain the high methylation level of the genome. Gene mismatch could be caused by the down regulation of MBD4 gene expression level. The results indicated that the abnormal expression of methylation factors could lead to abnormal methylation of the nuclear transfer embryos,which was the main cause of dysplasia or stillbirth.