抗人精浆蛋白单克隆抗体轻、重链可变区基因的克隆及序列分析

Cloning and sequcllcing of the VH/VL genes of antihuman γ-seminoprotein McAb

目的：获得鼠抗人精浆蛋白（γ－Sm）单抗可变区基因，为单链抗体的构建及表达奠定基础．方法：从分泌抗人精浆蛋白单克隆抗体（γ-Sm－McAb）的杂交瘤细胞系E4B7中提取总RNA，经反转录、PCR分离获得了γ－Sm－McAb轻链可变区（VL）基因和重链可变区（VH）基因，分别克隆人pUC19质粒中，用全自动荧光DNA序列分析仪对VH，VL基因序列进行了分析．结果：E4B7VH基因由358bp组成，编码119个氨基酸，E4B7VL基因由326bp组成，编码108个氨基酸；从推导出的氨基酸序列分析证实E4B7VH，VL序列均符合小鼠Ig可变区的氨基酸序列特征．结论：所克隆基因与Internet中基因文库的所有已知基因比较，未查到相同基因，与小鼠免疫球蛋白可变区基因同源性最高．

Aim:To clone the variable region gene of murineMcAb against γ-seminoprotein, for constructing the singlechain variable fragment. Methods:Total RNA was extractedfrom the hybridoma cell line E4B7, which secretes McAb against γ-seminoprotein, and subjected to reverse transcription. The VH and VL genes were amplified by PCR, clonedinto plasmid vector and analysed using auto DNA sequencer. Results: E4B7 VH consists of 358 hp encoding 119amino acid residues, E4B7 VL contains 326 hp encoding 108amino acid residues. The deduced amino acid sequences ofE4B7 VH/VL were in agreement with the characterization ofthe amino acid present in the mouse Ig variable region. Conclusion: The cloned sequences were both confirmed as newgenes by comparing with all known genes in GenBank in Internet. The sequences exhibite significant homology to previously reported antibody variable region genes.