摘要
目的:克隆并分析抗重组炭疽芽孢杆菌保护性抗原(rPA)单克隆抗体4B2、5E1和2A8轻、重链可变区基因。方法:从分泌抗rPA特异性单抗的杂交瘤细胞株4B2、5E1和2A8中提取总RNA;利用RT-PCR技术克隆抗rPA单抗4B2、5E1和2A8的VL、VH基因,并将其连入pMD18-T载体中进行测序分析。结果:4B2VL基因全长378bp,VH基因全长414bp;5E1VL基因全长420bp,VH基因全长426bp;2A8VL基因全长384bp,VH基因全长342bp。通过分析,这些基因均符合小鼠IgGV区基因的特征,含有4个框架区、3个抗原互补决定区,而且抗体特征性的半胱氨酸残基的数量和位置也正确。结论:克隆了抗rPA单抗4B2、5E1和2A8的VL、VH基因,为下一步构建多种形式的基因工程抗体奠定了良好的基础。
Objective： To clone and sequence VL, V. genes of monoclonal antibodies （mAb） against anthrax protective antigen. Methods： Total RNA was extracted from hybridoma cell 4B2, 5El and 2A8 secreting mAb against anthrax protective antigen, and VL and V. genes were amplified by RT-PCR, Then the PCR products were ligated to pMD18-T vector and the positive clones were analyzed. Results： The amplified VL and VH genes of 4B2 were 378 bp and 414 bp, respectively. The amplified VL and V. genes of 5El was 420 bp and 426 bp, respectively. The VL gene of 2A8 contained 384 bp and the V. gene of 2A8 contained 342 bp. All the sequences were analyzed by logging onto the sites of http：// www.ncbi.nlm.nih.gov/, http：//www.ebi,ac.uk/imgt/ and http：//www.cbs.dtu.dk/services/SignalP-2.O/. The results showed that these genes contained four FRs, three CDRs and two characteristic cysteine residues. Conclusion： The VL and V. genes of mAb 4B2, 5El and 2A8 against anthrax protective antigen were successfully cloned, which lay a good foundation for constructing a diversity of engineering antibodies.
出处
《生物技术通讯》
CAS
2007年第2期179-182,共4页
Letters in Biotechnology
基金
国家自然科学基金项目(30300016)
