抗心肌肌凝蛋白重链的单链抗体V_H和V_L基因的合成  

Synthesiaed Genes of VH and VL of Single-Chain Antibody of Human Cardiac Myosin Heavy Chain

作　　者：宋娜玲[1] 赵启仁[1] 张春明[1] 刘家慧[1] 刘洁[1] 褚丽萍[1] 王德芝[1] 贺欣[1] 王月英[1] 

机构地区：[1]中国医学科学院放射医学研究所天津分子核医学重点实验室,天津300192

出　　处：《中国辐射卫生》2012年第1期1-3,共3页Chinese Journal of Radiological Health

基　　金：国家自然科学基金项目(No.39970223);中国医学科学院放射医学研究所基金资助项目(No.SF1104)

摘　　要：目的为研制心肌显像剂—抗人心肌肌凝蛋白重链(HCMHC)的单链抗体,合成抗HCMHC单链抗体的VH和VL基因。方法用TRIZOL试剂从抗HCMHC McAb杂交瘤细胞提取总RNA,合成第一链cDNA,以这第一链cDNA为模板,用合成的特异引物、DNA聚合酶和四种单核苷酸,对重链可变区(VH)基因和轻链可变区(VL)基因进行PCR扩增。对各步骤的产物进行鉴定,并做了对RNA的稳定性与贮存温度的关系,MgCl2浓度和循环温度的最优化的选择。结果合成的第一链cDNA纯度达95%;扩增产物的琼脂糖凝胶电泳带呈单一明亮泳带,VH和VL基因分子量分别为340bp和320bp,与文献相一致。结论成功合成了VH和VL基因,为心肌显像及其显像剂抗HCMHC单链可变区抗体的研究打下了基础。Objective To synthesize genes of the heavy chain variable region （ VH ） and light chain variable region （ VL ） of single - chain antibody of human cardiac myosin heavy chain for the development of myocardial .imaging agents ： single - chain antibody of human cardiac myosin heavy chain. Methods To extracted total RNA of the anti - HCMHC McAb hybridoma using TRIzoL reagent, synthesize the first - strand eDNA, using this first - strand eDNA as template, with specific primers, DNA polymerase and four single nucleotides, amplifiy the genes of the heavy chain variable region （ Vn ） and light chain variable region （ VL ） by PCR. To identifiy products of each step and study relationship between RNA stability and storage temperature and optimize cycle selection temperature with MgCl2 concentration. Results The purity of the first - strand eDNA reached 95% , PCR products by agarose gel electrophoresis showed a single band with a bright swimming, molecular weight of Va and VL genes were 340bp and 320bp, which was consistent with literature reports. Conclusion Synthesized genes of VH and VL, laid the foundation for the development of myocardial imaging agents ： single - chain antibody of human cardiac myosin heavy chain.

关 键 词：心肌肌球蛋白重链 可变区抗体 VH基因 VL基因 

分 类 号：R817.4[医药卫生—影像医学与核医学]