重度子痫前期胎盘组织差异表达基因片段的克隆和初步鉴定

目的:寻找并克隆子痫前期胎盘组织差异表达的基因,为重度子痫前期发生、发展的分子机制提供线索。方法:应用荧光mRNA差异显示(FDD)技术,寻找重度子痫前期患者和正常孕妇胎盘组织基因表达的差异,对获得的差异基因片段进行PCR扩增、克隆及测序,应用BLAST软件在国际基因数据库中进行测序结果同源性的分析,应用Northern杂交验证部分差异表达基因。结果:子痫前期患者胎盘差异表达基因片段18条,表达上调的10条,下调的8条,在子痫前期患者胎盘中高表达的10个基因片段中,5个分别与丝-苏氨酸蛋白磷酸酶2A催化亚基β、ATP结合盒转运子G2、细胞色素氧化酶亚单位Ⅵc、δ-氨基乙酰丙酸脱水酶基因、空泡分选蛋白29基因高度同源,2个片段虽已有同源基因序列,但功能未知,3个未找到高度同源的序列,申请后获得了新的GenBank登录号CV998051、CV998052、CV998053;在子痫前期患者胎盘中低表达的8个基因片段中,5个分别与蛋白磷酸酶1调节亚单位2、核自身抗原、DNA聚合酶ε4基因及FAM13C1基因高度同源,3个无高度同源序列,申请获得GenBank登录号CV998054、CV998055、CV998057。Northern印迹杂交验证3个新登录基因片段的表达。结论:应用FDD技术从子痫前期患者胎盘组织中克隆出18个差异表达基因片段,为子痫前期相关基因的研究提供了新方向。

Cloning and identification of differentially expressed genes in the placenta from severe preeclamptic patientes

Objective:To provide clues for the molecular mechanism of preeclampsia by screening and cloning genes in placenta of preeclampsia.Methods:To search differentially expressed gene fragments in placenta between patients with severe preeclampsia and normal pregnant women by fluorescent differential display(FDD),and the differentially expressed genes were reamplified、cloned and sequenced,the sequenced results were undergone with homologous analysis through BLAST software in GenBank.To validate the expression of some genes by Northern hybridization.Results:Eighteen cDNA fragments was found.Ten of them were highly expressed in placentas of preeclamptic patients,five of them were respectively high homologous to human protein phosphatase 2A catalytic subunit-beta,ABC transporter G_2,delta-aminolevulinate dehydratase,cytochrome C oxidase subunit Ⅵc and vacuolar protein sorting 29,2 of them were homologous to genes which function was unknown so far,three of them had low identity with known genes and were submitted to GenBank,which were given new accession numbers of CV998051,CV998052,CV998053.8 of them were lowly expressed in placentas of preeclamptic patients,5 of them were respectively high homologous to human regulatory subunit 2 of protein phosphatase 1,nuclear autoantigen,DNA-directed polymerase epsilon 4 and family with sequence similarity 13,member C1,3 of them were new cDNA sequences and submitted to GenBank,which were obtained accession numbers of CV998054,CV998055 and CV998057.The expression of three noval gene fragments were verified by Northern blot.Conclusion:Eighteen differentially expressed fragments in the placenta of patients with preeclampsia are cloned by FDD,these genes provide new directions for the study of the pathogenesis of preeclampsia.