日本血吸虫单克隆抗抗独特型抗体NP48可变区基因的克隆及序列分析

CLONING AND SEQUENCE ANALYSIS OF VARIABLE REGION GENE OF MONOCLONAL ANTI-ANTI-IDIOTYPIC ANTIBODY NP48 OF SCHISTOSOMA JAPONICUM

目的 克隆日本血吸虫单克隆 anti- anti- id NP4 8轻链及重链可变区基因并分析其序列。方法 从杂交瘤细胞 NP4 8提取总 RNA、RT- PCR扩增目的基因片段 ,分别克隆于 p MD18- T载体 ,测序并作核苷酸序列分析。结果 VL 基因全长 336 bp,属鼠免疫球蛋白κ链第 亚类 ,由种系基因he2 4与 Jκ4 重排而来。该 VL基因序列已被 Gene Bank收录 (accession No.AY 30 90 10 )。 VH 基因全长35 4 bp,属鼠免疫球蛋白重链第 亚类 ,由种系 J5 5 8.4 7、Dsp2 .2和 JH4 基因重排而来。该 VH 基因序列已被 Gene Bank收录 (accession No.AY312 4 33)。结论 序列比对分析表明该 VL、VH基因为日本血吸虫单克隆 anti- anti- id NP4 8可变区基因。

Objective To clone the variable region genes of light and heavy chains (V L and V H) of monoclonal anti-anti-idiotypic antibody NP48 of Schistosoma japonicum, and to analyze the nucleotide sequences of the V L and V H genes. [WT5”HZ]Methods The hybridoma cells secreting monoclonal anti-anti-idiotypic antibody NP48 of Schistosoma japonicum[WT5”BZ] were cultured and their total RNA was extracted. The desired V L and V H fragments were amplified by RT-PCR from the total RNA. The PCR products were then cloned into pMD18-T vector and sequenced. The V L and V H gene were compared with the Genebank and published mouse variable region genes. Results It was proved that the full-length V L gene was 336 bp, and a member of mouse Ig κ chain subgroup Ⅱ and originated from rearrangement of germ line he24 and Jκ 4 genes. The V L gene sequence was registered by the Genebank (accession No. AY309010). It was proved that the full-length V H gene was 354 bp, and a member of mouse Ig heavy chain subgroup Ⅱ and originated from rearrangement of germ line J558.47, Dsp2.2 and J H4 genes. The V H gene sequence has been registered by the Genebank (accession No. AY312433). Conclusion The obtained V L and V H genes are variable region genes of monoclonal anti-anti-idiotypic antibody NP48 of Schistosoma japonicum.