阴道毛滴虫衰老生物学标志分子β.DOCVIP

临床医学论文-阴道毛滴虫衰老生物学标志分子β_半乳糖苷酶基因克隆和蛋白表达 【摘要】? 目的:克隆并分析阴道毛滴虫(Trichomonas vaginalis, Tv)衰老生物学标志分子β_半乳糖苷酶(β_gal)同源基因，表达β_GAL重组蛋白。方法:从Tv cDNA表达文库中分离获得Tv　β_gal同源基因的cDNA克隆，测序和序列分析。将cDNA部分片段亚克隆到原核表达载体(pQE_80L)，转化宿主菌E.coli SG13009，异丙基_β_D_硫代半乳糖苷诱导表达，聚丙烯酰胺凝胶电泳分析鉴定。结果:在Tv cDNA文库中获得1株2445bp的cDNA克隆。序列分析表明，该序列开放读码框有2415bp，推测肽链具有804个氨基酸,等电点为54；该肽链与多种来源的β_GAL同源蛋白均具有较高同源性。用PCR扩增了该cDNA序列中的2277bp片段，构建了pQE_80L/Tv　β_gal,所表达重组蛋白产物的相对分子质量为82ku。结论:推测该克隆是Tv　β_gal的同源基因，Tv　β_gal基因可以在E.coli中得到表达，为进一步研究其细胞内定位和蛋白功能奠定了基础。 【关键词】? 阴道毛滴虫 β_半乳糖苷酶 cDNA克隆 ??? 2Heyuan Entry_Exit Inspection and Quarantine Bureau, Heyuan 517000, China)［Abstract］Objective：To clone and analyze Trichomonas vaginalis(Tv)β_galactosidase(Tv_β_gal)homology gene , and to express the recombinant protein. Methods：One cDNA clone, with high homology of β_gal, was isolated from Tv cDNA library. The cDNA was sequenced and analyzed. Then a fragment of the cDNA was cloned to the expression vector pQE_80L. E.coli SG13009 cell was transformed by the recombinant plasmid. The expression of protein was induced by isopropyithio_β_D_galactoside. Sodium dodecyl sulfate_polya_crylamide gel electrophoresis analysis was performed to detect the recombinant protein. Results：One cDNA clone with a length of 2 445 bp was isolated. The cDNA clone had an open reading frame with 2 415 bp. The deduced amino acid sequence contains 804 residues and its isoelectric point is 54. The cDNA clone was high homology to the β_gal homologus of different species. A 2 277 bp fragment of the cDNA was amplified by PCR and therecombinant expression plasmid(pQE_80L/Tv β_gal)was constructed. The fusion protein of Tv β_GAL(about 82 ku)was expressed. Conclusion：The cDNA clone isolated belongs to β_gal homolog. Tv β_gal is expressed well in E.coli, which will be helpful for the further study on the location and function of Tv β_gal in Tv. ??? ［Key Words］Trichomonas vaginalis; β_galactosidase; cDNA clone 　　β_半乳糖苷酶(β_GAL)，广泛存在于各种动物、植物及微生物中。早期只是利用该酶水解乳糖的性质来降低牛乳中的乳糖含量,改善人类对牛乳乳糖的不