冷冻保存对猪耳成纤维细胞H19和IGF2R基因DNA甲基化和基因表达的影响  

Effects of DNA Methylation Status of H19, IGF2R and Genes Expression of Porcine Ear Skin Fibroblasts Induced by Cryopreservation

作　　者：张宏丽 陈晓 戴建军[1,2,3] 姚惠娟 张树山[1,2,3] 张德福 ZHANG Hong-li;CHEN Xiao;DAI Jian-jun;YAO Hui-juan;ZHANG Shu-shan;ZHANG De-fu(Animal Husbandry and Veterinary Research Institute,Shanghai Academy of Agricultural Science,Shanghai 201106,China;Division of Animal Genetic Engineering,Shanghai Key Laboratory of Agri-Genetics and Breeding,Shanghai 201106,China;Shanghai Engineering Research Center of Breeding Pig,Shanghai 201106,China)

机构地区：[1]上海市农业科学院畜牧兽医研究所,上海201106 [2]上海市农业遗传育种重点实验室动物遗传工程研究室,上海201106 [3]上海种猪工程技术研究中心,上海201302

出　　处：《中国畜牧杂志》2018年第7期86-90,共5页Chinese Journal of Animal Science

基　　金：国家自然科学基金(31372315);上海市农业科学院攀高计划(PG171);嘉定区农业和社会事业项目(梅山猪分子(基因)保种技术研究;2016004)

摘　　要：为探索冷冻保存对猪体细胞H19和IGF2R基因DNA甲基化的影响,本研究以梅山猪耳成纤维细胞为材料,采用亚硫酸氢盐测序和RT-PCR技术,检测了新鲜和冷冻保存细胞中H19和IGF2R基因的DNA甲基化状态和表达量,并对甲基化相关基因的表达水平进行分析。结果表明:冷冻组中H19基因DMR1和DMR3的甲基化率极显著高于新鲜组(P<0.01),H19基因DMR2表现为去甲基化,极显著低于新鲜组中的甲基化率(P<0.01),且该基因表达量极显著高于新鲜组(P<0.01);冷冻组IGF2R基因DMR2表现为超甲基化,冷冻组极显著高于新鲜组中的甲基化率(P<0.01),但IGF2R基因表达量无显著差异(P>0.05);冷冻组中DNMT3A和DNMT1的基因表达量极显著高于新鲜组(P<0.01),DNMT3B基因表达量则无显著差异(P>0.05)。本实验条件下,冷冻保存影响了H19和IGF2R基因甲基化控制区的DNA甲基化状态,从而影响了该基因的表达水平。In order to explore the effects of methylation status of H19 and IGF2R genes in porcine somatic cells induced by cryopreservation,DNA methylation status and expression levels of H19,IGF2R in Meishan pig ear fibroblast cells before and after cryopreservation were detected with bisulfite sequencing and RT-PCR technology.The expression levels of methylation related genes were also detected and analyzed.The results showed that the methylation rates of H19 DMR1 and DMR3 in cryopreserved cells were significantly higher than those in fresh cells(P<0.01).H19 DMR2 was demethylation,and the methylation rates of H19 DMR2 was significantly lower than that in fresh cells(P<0.01),and expression level of H19 gene was significantly higher than that in the fresh cells(P<0.01).The IGF2R DMR2 was hypermethylation,and its methylation rates was significantly higher than that in the fresh cells(P<0.01),but there was no significant difference in the expression level of IGF2R DMR2 between fresh and cryopreserved cells(P>0.05).The expression levels of DNMT3A and DNMT1 in cryopreserved cells were significantly higher than those in fresh cells(P<0.01),and there was no significant difference in the expression level of DNMT3B(P>0.05)between two groups.The results indicated that the cryopreservation could affect the methylation status of H19 and IGF2R DMR,thus changing their gene expression levels.

关 键 词：梅山猪 耳成纤维细胞 冷冻保存 DNA甲基化 基因表达 

分 类 号：S828.2[农业科学—畜牧学]