长白山白眉蝮蛇乌苏里亚种类凝血酶(TLE)基因克隆及分析  

Cloning and Bioinformation Analysis of a Novel Thrombin-like Enzyme Gene from Gloydius ussuriensis

作　　者：薛雁[1] 孙东[1] 于翀[1] 宁静[1] 崔亮亮[1] 石皎[1] 

机构地区：[1]沈阳守正生物技术有限公司,辽宁沈阳110171

出　　处：《蛇志》2011年第4期341-344,360,共5页Journal of Snake

摘　　要：目的为了获得长白山白眉蝮蛇乌苏里亚种类凝血酶基因。方法根据Gene Bank白眉类凝血酶cDNA 5'和3'保守序列设计了引物,通过RT-PCR从白眉蝮蛇乌苏里亚种毒腺Total RNA中扩增得到1条长714bp的特异cDNA片段,将该cDNA片段重组到Simple Tvector,转化进E.coli JM 109competent cell,阳性克隆委托生物公司测序,利用生物信息学方法对测序结果进行分析。结果该特异性片段与蛇毒类凝血酶同源性为95%,它为一个开发阅读框架,其编码的蛋白质序列与其他蛇毒类凝血酶序列同源性为94%,与其他蛇毒类凝血亲缘关系非常近。结论本实验获得了一种新型白眉蝮蛇乌苏里亚种类凝血酶基因。Objective To obtain the thrombin-like enzyme gene of snake venoms. Methods Two primers were designed according to the snake venom thrombin-like enzyme （TI.E） highly conserved regions of 5＂and 3＂. Primer 1 is 5＂-CGCGAATTCATGGTCATTGGAGGT- GTT-3＂ and Primer 2 is 5-ATACTCGAGTCACTGGGGGCAGGTCG-3＂ . Total RNA was pre- pared from the venom gland of Gloydius ussuriensis and RT-PCR was conducted to amplify the TLE. A 716 bp DNA fragment was obtained,inserted into the pMD18-T vector and transferred into E. coli JM 109 competent cell, then identified by restriction enzyme and sequencing Re- sults The results showed that the cDNA we got shared great sequence homology （95%） with the published snake TI.E eDNA sequence,the amino acid sequence encoded by this eDNA was 237,which shared 94% homology with Gene Bank, it has very high homology with other snakes. Conclusion A novel eDNA encoding snake TLE was got by RT-PCR.

关 键 词：类凝血酶 白眉蝮蛇 CDNA序列 

分 类 号：R996.3[医药卫生—毒理学] R977.3[医药卫生—药学]