摘要
目的 :克隆并表达人可溶性增殖诱导配体 (sAPRIL ,即人APRIL10 5 2 50 ) ,为探索其在多种肿瘤细胞的增殖和存活以及促肿瘤形成中的作用奠定基础。方法 :从GENBANK中查找人APRIL蛋白 (编号 :0 75 88)序列 ,取其部分胞外 (APRIL10 5 2 50 )序列设计引物 ,用RT PCR从扁桃体总RNA中扩增出人APRIL10 5 2 50 基因 ,测序后将克隆载体经酶切并构建表达载体 ,在大肠杆菌中表达 ,并纯化蛋白。结果 :经克隆测序后进行同源比较 ,证实所克隆的基因即为人APRIL10 5 2 50 基因。在大肠杆菌中表达量达 43 6% ,获得纯化蛋白。结论 :成功克隆与表达、纯化了人APRIL10 5 2 50 基因 。
Objective:To obtain human soluble APRIL cDNA and stable expression on E.coli DH5α.Methods: Screening the human APRIL protein sequence from GenBank(No:07588),designing the primer according to part of its extracelluar protein sequence.The total RNA,extracted from human tonsil,was amplified into the sequence of cDNA encoding sAPRIL by RT PCR and cloned into vector pUC18 for sequencing.Next,the identified cDNA was cloned into the new type of prokaryotic expression vector pQE 80L.Transmited the combined plasmids into E.coli DH5α and purifieded the protein by Ni 2+ .Results: The sAPRIL molecule was successfully expressed on E.coli DH5α and the protein was purifieded.Conclusion:The sAPRIL was expressed stably on E.coli DH5α.These works may provide evidence for the further study of the sAPRIL protein.
出处
《中国生物工程杂志》
CAS
CSCD
2004年第10期38-41,共4页
China Biotechnology
