乙肝病毒感染动物模型喜马拉雅旱獭IL-15分子的克隆及序列分析

摘要

Objective To clone and analyze the sequence of interleukin-15 (IL-15) in Marmota himanayana ,the newly es-tablished animal model for the study of hepatitis B virus (HBV) infection.Methods Total RNA was isolated from spleen tissues of Marmota himanayana.cDNA fragments of IL-15 in Marmota himanayana was amplified by reverse transcription polymerase chain reaction (RT-PCR) and cloned into the vector pMD18-T.The recombinant plasmids were identified by restricted enzyme digestion and sequenced commercially.The homology of IL-15 in Marmota himanayana to other mammal animals ,its affinity to other species and the protein structure of IL-15 were analyzed by using some software packages.Results Sequence analysis in-dicated that cDNA fragments (489 bp) coding IL-15 (162 amino acids) was obtained.Homology and phylogenetic analysis indi-cated that IL-15 in Marmota himanayana had the highest homology to that in woodchuck and the lowest homology to that in rat.The homology modeling revealed that the 3D structure of IL-15 protein in Marmota himanayana was similar to that of woodchuck.Conclusion The complete sequences of IL-15 in Marmota himanayana was obtained ,which provides a basis for further studying the Chinese Marmot animal model.%目的：对新近建立的乙肝病毒感染动物模型喜马拉雅旱獭的白细胞介素-15（IL-15）分子进行克隆和序列分析。方法抽提喜马拉雅旱獭脾脏总 RNA ，RT-PCR扩增旱獭IL-15 cDNA序列，克隆至pMD18-T载体，制备重组质粒酶切、基因测序鉴定。利用软件分析旱獭IL-15与其他哺乳动物IL-15的同源性、种系亲缘关系，并预测蛋白结构。结果 RT-PCR扩增获得大小为489 bp的序列，编码162个氨基酸。旱獭IL-15和土拨鼠IL-15同源性最高，和大鼠同源性最低。种系进化树结果提示旱獭IL-15与土拨鼠IL-15亲缘关系最近。旱獭IL-15蛋白构象与土拨鼠IL-15空间构象非常相似。结论克隆了喜马拉雅旱獭IL-15完整编码序列，为更好地利用新型乙肝病毒感染动物模型喜马拉雅旱獭奠定了基础。