山羊克隆胎儿不同组织中H19基因CpG岛甲基化模式及表达量检测

CpG island methylation patterns and expressions of H19 gene in cloned fetus of goat

摘要:

表观重编程异常是核移植胚胎发育异常的重要原因。为了研究克隆山羊胎儿不同组织中H19基因CpG岛甲基化水平和相对表达量，本实验运用亚硫酸盐法和荧光实时定量PCR法分别检测了死亡克隆山羊胎儿和同期普通山羊胎儿 (对照组) 肝脏、胎盘、肾脏、肺脏和心脏组织中H19基因CpG岛甲基化水平和mRNA的相对表达量。结果表明，克隆山羊胎儿胎盘组织中H19基因第5个CpG岛的甲基化水平显著高于对照组 (70％ vs 49.41％，P＜0.05)，H19基因相对表达量显著低于对照组 (883.3 vs 1264.5，P＜0.05)；肺脏组织甲基化水平显著低于对照组 (63.53％ vs 88.24％，P＜0.05)，相对表达量显著高于对照组 (1003.4 vs 515.5，P＜0.05)；其他各组差异不显著 (P＞0.05)。结果说明，H19基因在克隆山羊胎儿部分组织中DNA甲基化重编程异常，而且这种异常影响H19基因的正常表达，这也可能是导致克隆动物死亡的重要因素之一。

Abstract:

The aberrant epigenetic reprogramme is an important cause for abnormal development of nuclear transfer embryos. The objective of this study was to investigate the CpG island methylation profiles and relative expression levels of H19 gene in different tissues of cloned goat fetus. We detected liver, placenta, kidney, lung and heart in the dead cloned goat fetus and the age-matched normal goat fetus (control) by using bisulfite sequencing and real time PCR. Results indicated that methylation levels of the fifth CpG island of H19 gene in dead cloned goat fetus was significant high compared with that in the control in placenta (70% vs 49.41%, P < 0.05), and relative expression levels of H19 gene was significant low compared with that in the control (883.3 vs 1264.5, P < 0.05). Reversely, the methylation levels was significant low compared with that in the control in lung (63.53% vs 88.24%, P < 0.05), and relative expression levels was significant high compared with that in the control (1003.4 vs 515.5, P < 0.05). The differences of others groups were insignificant (P > 0.05). Results showed the abnormal DNA methylation proflies of H19 gene occurred in some tissues of cloned goat fetus, which affected normal expression levels of H19 gene, indicating that aberrant DNA methylation reprogramme may be one of the important factors for the death of cloned animals.