人肝脏特异性miR-122表达载体的构建及鉴定.PDF

生 物 工 程 学 报 Chin J Biotech 2009, April 25; 25(4): 587-590 journals.im.ac.cn Chinese Journal of Biotechnology ISSN 1000-3061 cjb@im.ac.cn © 2009 Institute of Microbiology, CAS CSM, All rights reserved 医学与免疫生物技术 miR-122 1 2 黄增红 , 刘长梅 1 , 100193 2 , 100101 : 人肝脏特异性 miRNA-122 是肝脏中表达丰度最高的 miRNA 。为研究该miR-122 的生物学功能, 从 HepG2 细 胞基因组中用PCR 的方法扩增了miR-122 的前体, 构建了miR-122 的表达载体pLMP-miR-122 。pLMP-miR-122 质粒转 染人正常肝细胞系L-O2 和肝癌细胞系HepG2 后, 细胞内成熟miRNA-122 的表达量显著增加。该质粒与HBV1.3 共转 染HepG2 细胞72 h 后, HBV 的HBs 和HBe 蛋白水平的表达量均下降, 说明miRNA-122 参与了HBV 基因的复制和表 达的调控, 为进一步研究miRNA-122 的功能和其他一些肝病如HCC 的调控机制打下基础。 : miRNA, miR-122, pLMP 质粒, HBV Construction and identification of the human liver-specific miR-122 expression vector 1 2 Zenghong Huang , and Changmei Liu 1 College of Biological Science, China Agricultural University, Beijing 100193, China 2 Laboratory of Molecular Virus , Institute of Microbiology , Chinese Academy of Sciences, Beijing 100101, China Abstract: miR-122 is the most abundant miRNA in adult human liver. To study the functions of miR-122 in liver disease, we amplified the precursor of human miR-122 gene by polymerase chain reaction (PCR) from the HepG2 genomic DNA, and then constructed miR-122 expression vector pLMP-miR-122. pLMP-miR-122 could overexpress mature miR-122 when human normal liver cells L-O2 and the hepatoma cells HepG2 were transfected with it. When HepG2 cells were co-transfected with HBV1.3 and pLMP-miR-122, we found that miR-122 could down-regulate the