血管内皮细胞乙酰胆碱作用靶标相关新基因的电子克隆

血管内皮细胞乙酰胆碱作用靶标(ETA)作为特异性血管内皮细胞药物作用新靶点具有良好的抗动脉粥样硬化应用前景。通过培养前后血管内皮细胞表达差异基因的筛选，有可能分离获得ETA或其相关基因。在利用抑制削减杂交技术获得14个牛未知基因片段基础上，采用电子克隆途径，使6个未知基因片段得以延伸，并在GenBank的非冗余基因库中找到了对应或同源基因，其中3个延伸出了全长阅读框，另外3个延伸出部分长度；对于延伸出全长阅读框的基因，设计特异性引物进行PCR扩增，验证了电子延伸的可信性，同时也克隆了整个阅读框架区。

The electronic cloning of ETA related unknown genes

Endothelial target for acetylcholine(ETA)plays an important role in preventing atherosclerosis,and it may be a unique molecular target of the novel drugs against atherosclerosis.Study of differential expression genes between fresh and cultured vascular endothelial cell(VEC)will help us isolate ETA or its related genes.In former work,suppression sub-tractive hybridization technique was employed to construct the differential expression gene segments library,from which14unknown down-regulated bovine gene segments were identified.In this study,six ESTs lengths were extended and three of them obtained full-length ORF while the other three got partial-length ORF through electronic cloning.The corresponding or homology genes of the six extended ESTs have been found in GenBank.Specific primers for the three full-length ORF genes were designed and PCR was used to validate the reliability of electronic cloning,and the full-length ORF of these three genes were cloned as well.