牛KLF14和PDE10A基因的印记鉴定和甲基化分析  

Imprinting Identification and Methylation Analysis of Bovine(Bos taurus)KLF14 and PDE10A Genes

作　　者：董艳秋 靳兰杰 杨欣怡 李冬杰[2] 霍浩楠 苏红 李世杰[1] DONG Yan-Qiu;JIN Lan-Jie;YANG Xin-Yi;LI Dong-Jie;HUO Hao-Nan;SU Hong;LI Shi-Jie(College of Life Sciences,Hebei Agricultural University,Baoding 071000,China;College of Bioscience and Bioengineering,Hebei University of Science and Technology,Shijiazhuang 050018,China;Baoding University of Technology,Baoding 071000,China)

机构地区：[1]河北农业大学生命科学学院,保定071000 [2]河北科技大学生物科学与工程学院,石家庄050018 [3]保定理工学院,保定071000

出　　处：《农业生物技术学报》2023年第4期776-783,共8页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金(31372312);河北省自然科学基金(C2020204004);保定市科技局项目(2041ZF202)。

摘　　要：在哺乳动物中,印记基因与胎盘发育和胎儿生长密切相关。印记基因的表达变化会通过各种生理反应和信号通路导致哺乳动物生长发育异常并引发相关疾病。KLF14(Kruppel like factor 14)和磷酸二酯酶10a(phosphodiesterase 10a,PDE10A)基因在人(Homo sapiens)和小鼠(Mus musculus)中被鉴定为印记基因。目前,关于KLF14和PDE10A基因在牛(Bos taurus)中的印记研究还未见报道。本研究首先应用RTPCR方法分析KLF14和PDE10A基因在牛心脏、肝脏、脾脏、肺脏、肾脏、大脑和胎盘中的表达,发现KLF14基因只在胎盘中表达;而PDE10A基因在被检测组织中均有表达。进一步应用基于SNP的RT-PCR产物直接测序方法,分析KLF14和PDE10A的等位基因表达状态。结果显示,KLF14基因在胎盘中为单等位基因表达,通过对其双亲的基因型进行分析,确定KLF14基因在牛胎盘中为母源印记基因;而PDE10A基因在胎盘和其他组织中均呈现双等位基因表达,为非印记基因。应用亚硫酸盐测序方法分析KLF14基因启动子区的甲基化状态,发现其在牛胎盘和精子中均为轻甲基化,不存在差异甲基化区,说明KLF14基因启动子区的甲基化修饰不参与调控KLF14基因的印记表达。上述结果表明牛KLF14基因为胎盘特异的母源印记基因,PDE10A基因为非印记基因。本研究为深入探讨KLF14和PDE10A基因功能提供参考依据。In mammals,imprinted genes play key role in placental development and fetal growth.Changes in the expression of imprinted genes can lead to abnormal growth and development in mammals and cause diseases through various physiological responses and signaling pathways.KLF14(Kruppel like factor 14)and phosphodiesterase 10a(PDE10A)genes were identified as imprinted genes in human(Homo sapiens)and mouse(Mus musculus).At present,the imprinting studies on KLF14 and PDE10A in cattle(Bos taurus)have not been reported.In this study,the expression of KLF14 and PDE10A genes in bovine heart,liver,spleen,lung,kidney,brain and placenta were firstly analyzed by RT-PCR.The results showed that KLF14 gene was only detected in the placenta;PDE10A gene was expressed in all tested tissues.The allelic expression status of KLF14 and PDE10A genes were further analyzed by direct sequencing of RT-PCR products based on SNP method.KLF14 gene showed monoallelic expression in the placenta.By analyzing the genotypes of parental genomic DNA,KLF14 gene was identifiied to be a maternally imprinted gene in bovine placenta.PDE10A gene showed biallelic expression in placenta ant other tissues.PDE10A gene was non-imprinted in bovine.The methylation status of the promoter region of KLF14 gene was analyzed by sulfite sequencing method,hypomethylation were found in both bovine placenta and sperm,and there was no differentially methylated region,which indicated that the methylation modification of the promoter region of KLF14 gene might not be involved in regulating the imprinted expression of KLF14 gene.The above results suggest that bovine KLF14gene is a placenta-specific maternally imprinted gene and bovine PDE10A gene is non-imprinted gene.This study provides a reference for further study of the function of the bovine KLF14 and PED10A.

关 键 词：牛 基因组印记 DNA甲基化 KLF14(Kruppel like factor 14) 磷酸二酯酶10a(PDE10A) 

分 类 号：S823[农业科学—畜牧学] Q78[农业科学—畜牧兽医]