摘要
【目的】在家蚕基因表达谱研究中鉴定了近4万个特异的SAGE标签,为了研究这些标签所对应基因的功能,选择了其中一个SAGE标签,探索这个SAGE标签对应的基因序列,表达特征以及相关的功能。【方法】在SAGE结合GLGI结果的基础上,利用5’RACE获得基因的全长,根据不同物种间该基因的同源性来构建系统发生树,通过PCR检测该基因在家蚕不同时期、不同组织中的表达特征,并在原核表达系统中表达该基因。【结果】从cDNA中获得了家蚕TCTP基因的全长序列,构建了不同物种间TCTP的系统发生树,PCR结果表明TCTP基因在家蚕的各个时期、各个组织中都稳定的表达,并且成功的在原核表达系统中表达了该基因。【结论】从cDNA中获得了TCTP基因的全长序列,并且成功在原核系统中表达,证明了通过笔者的实验方法能够获得新的基因;系统发生树显示亲缘性越近的物种TCTP间的同源性越高;家蚕TCTP基因在家蚕生命的各个时期、各个组织都有重要的生物学功能;这些工作为进一步研究其具体功能奠定了基础。
[Objective] Nearly 40 000 unique SAGE Tags of Bombyx mori were identified through serial analysis of gene expression technique. In order to understand the function of these Tags, one of them was selected to get the knowledge of the gene sequence and its expression profiles. [Method] Firstly, the SAGE Tag was elongated through GLGI to several hundreds base pair of EST; Secondly, through 3＇RACE and 5＇RACE, the full length gene sequence was obtained. Thirdly, the homology of the Tag corresponding sequence was analyzed with other species and a primary phylogenetic tree was constructed. In addition, the gene from various stage or tissue of Bombyx mori was amplified to achieve the gene expression profile in silkworm. Furthermore, the gene was cloned into E. coli to test its expression. [Result] The full length cDNA sequence with 794 bp of silkworm TCTP gene was obtained and the recombination protein was obtained through E. coli expression system. The RT-PCR expression tests indicated that the TCTP gene expressed in various tissues ofBombyx mori at different stages. [Conclusion] TCTP gene obtained from SAGE Tag is one of the important genes in silkworm. It does not have specific expression profiles at different stages. The results show that it is possible to get genes from SAGE tags for gene identification and expression study.
出处
《中国农业科学》
CAS
CSCD
北大核心
2007年第8期1809-1816,共8页
Scientia Agricultura Sinica
基金
国家"973"计划项目(2005CB121000)
自然科学基金项目(30500340)
关键词
家蚕
TCTP
同源性
系统发生树
原核表达
Bombyx mori
TCTP
Homology
Phylogenetic clustering
Prokaryotic expression
