9．1C3单克隆抗体轻、重链可变区基因克隆及序列分析

Cloning and Sequencing of the V_H/V_L Genes of 9. 1C3 McAb

９．１Ｃ３分子是一种参与ＮＫ、巨噬细胞及ＬＡＫ细胞杀伤过程的重要细胞表面分子，主要表达于外周血单个核细胞表面。本文运用分子生物学技术，从分泌９．１Ｃ３ＭＣＡｂ的杂交瘤细胞中提取总ＲＮＡ，经反转录、ＰＣＲ分另１１分离了９．１Ｃ３ＭｃＡｂ轻链可变区（ＶＬ）基因及重链可变区（ＶＨ）基因，并用全自动荧光ＤＮＡ序列分析仪对９．１Ｃ３ＶＨ、ＶＬ基因序列进行了分析。结果表明：９．１Ｃ３ＶＨ基因为３５１ｂＰ，编码１１７ａａ残基，９．１３ＶＬ为３２４ｂＰ，编码１０８ａａ残基；从推导出的氨基酸序列分析证实９．１Ｃ３ＶＨ、ＶＬ序列均符合小鼠ｌｇ可变区的氨基酸序列特征；根据Ｋａｂｂａｔ分类体系，９．１Ｃ３ＶＨ隶属Ｉｇ重链第Ⅱ（Ｃ）亚组，９．１Ｃ３ＶＬ隶属小鼠ＩｇＫ链第Ⅴ亚组。９．１Ｃ３ＶＨ、ＶＬ基因的克隆成功为其单链抗体的构建、表达奠定了良好的物质基础。

C3 molecules are expressed on Peripheral blood mononuclear cells and played an important role in Cytotoxic effects mediated by NK.M and LAK cells. In this paper, the VH and VL genesfrom hybridoma cell line 9. 1C3 secreting antibody against this molecule were amplified and sequencedby PCR. The VH and VL genes of 9. 1C3 McAb were analysed using auto DNA sequencer. The resultsshowed that 9. 1C3 VH consists of 351 bp encoding 1 17 amino acid residues, and 9. 1C3 VL contains324 hp encoding 108 amino acid residues.The deduced amino acid sequences of 9.1C3 VH/VL were inagreement with the characterization of the amino acid present in the mouse Ig variable region.According to kabbat classified method, 9. 1C3 VH and VL genes belongto the mouse ig heavy subgroup Ⅱ(C) and K chain subgroup Ⅴrespectively. The success of cloning of 9.1C3 VH/VL genes laya goodfundation for the construction and expression of 9. 1C3 single chain variable fragment.