人肝脏高凝状态相关基因的筛选  

Screening of Prothrombotic State-related Genes from Human Hepatic cDNA Library

作　　者：崔晓燕[1] 汤慧[1] 方定志[1] 刘秉文[1] 

机构地区：[1]四川大学华西基础医学与法医学院生物化学与分子生物学教研室,成都610041

出　　处：《四川大学学报（医学版）》2005年第5期605-608,共4页Journal of Sichuan University(Medical Sciences)

基　　金：国家973计划(项目编号G2000056900)资助

摘　　要：目的筛选人肝脏高凝状态(PTS)相关基因的全长cDNA序列。方法采用差异筛选法对本室建立的PTS大鼠肝脏差异表达基因消减cDNA文库进行筛选;以获得的差异cDNA克隆为模板,采用PCR方法扩增目的片段;以PCR产物作探针,用噬菌斑原位杂交法筛选人肝脏cDNA文库,经初筛、二筛和三筛后,将获得的阳性克隆转入E.coliBM25.8,使λTripIEx环化成质粒pTripIEx,经酶切鉴定后进行DNA测序。结果从人肝脏cDNA文库中筛选获得4条全长cDNA序列,经序列测定及生物信息学分析发现,其中3条的表达产物分别为纤维蛋白原、纤维蛋白原相关蛋白、10号染色体开放阅读框架104mRNA;另1条与氨基甲酰磷酸合成酶1同源。结论以PTS大鼠肝脏差异表达基因cDNA为探针,从人肝脏cDNA文库中筛选获得4条PTS相关cDNA全长序列。Objective To screen prothrombotic state-related cDNA sequences from human hepatic cDNA library. Methods The subtracted cDNA library for differentially expressed genes in rat liver of prothrombotic state （PTS） was constructed by suppression subtractive hybridization. Positive clones were identified by differential screening. The target DNA sequences of positive cDNA clones of differentially expressed genes from rat liver of PTS were amplified by PCR. The PCR products were used as probes to screen a human hepatic cDNA library. After the first, second and third screening, the transduction of a positive λTripIEx lysate into E.coli strain BM25. 8 promoted the circularization of pTripIEx. The positive circularized plasmids were identified by double enzyme digestion. The cDNA fragments of the positive plasmids were sequenced and analyzed by bioinformatics （blastn）. Results Four PTS-related cDNA sequences were identified from human hepatic cDNA library. For 3 of them, their products of expression were respectively fibrinogen, LFIRE1, and chromosome 10 open reading frame 104 mRNA. The fourth sequence had homology with carbamoyl-phosphate synthetase 1 gene. Conclusion Four PTS-related cDNA sequences have been identified from human hepatic cDNA library.

关 键 词：差异表达基因 CDNA文库 高凝状态 肝脏 

分 类 号：R575[医药卫生—消化系统]