Cui Xiao-Yan, Tang Hui, Fang Ding-Zhi, Liu Bing-Wen
Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2005 Sep;36(5):605-8.
To screen prothrombotic state-related cDNA sequences from human hepatic cDNA library.
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 lambdaTripIEx 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).
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.
Four PTS-related cDNA sequences have been identified from human hepatic cDNA library.
从人肝脏cDNA文库中筛选与血栓前状态相关的cDNA序列。
采用抑制性消减杂交技术构建血栓前状态（PTS）大鼠肝脏差异表达基因的消减cDNA文库。通过差异筛选鉴定阳性克隆。对PTS大鼠肝脏差异表达基因阳性cDNA克隆的目标DNA序列进行PCR扩增。PCR产物用作探针筛选人肝脏cDNA文库。经过第一、第二和第三次筛选后，将阳性λTripIEx裂解物转导至大肠杆菌菌株BM25.8中促进pTripIEx环化。通过双酶切鉴定阳性环化质粒。对阳性质粒的cDNA片段进行测序，并通过生物信息学（blastn）分析。
从人肝脏cDNA文库中鉴定出4条与PTS相关的cDNA序列。其中3条的表达产物分别为纤维蛋白原、LFIRE1和10号染色体开放阅读框104 mRNA。第四条序列与氨甲酰磷酸合成酶1基因具有同源性。
从人肝脏cDNA文库中鉴定出4条与PTS相关的cDNA序列。
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