抗人卵巢癌单克隆抗体COC183-B_2重链可变区基因的体外扩增、克隆及序列分析

AMPLIFICATION,CLONING AND SEQUENCE ANALYSIS OF THEHEAVY CHAIN VARIABLE REGION GENE OF MONOCLONALANTIBODY COC183B2 AGAINST HUMAN OVARIAN CARCINOMA

目的：分离抗人卵巢癌单克隆抗体ＣＯＣ１８３Ｂ２重链可变区（ＶＨ）基因并测定其序列。方法：用反转录ＰＣＲ扩增抗人卵巢癌单抗ＣＯＣ１８３Ｂ２ＶＨ基因，将其克隆入ＰＵＣ１９载体，重组子用Ｓａｎｇｅｒ’ｓ双脱氧链终止法测定序列，将序列与ＧｅｎｅＢａｎｋ及已发表的抗体序列比较。结果：ＶＨ基因全长３５４ｂｐ，属鼠免疫球蛋白重链混杂亚类（ｓｕｂｇｒｏｕｐｍｉｓｃｅｌａｎｅｏｕｓ），由种系基因的ＶＨ，Ｄｓｐ２．５及ＭＵＳＪＨ４重排而来。该ＶＨ基因序列已被ＧｅｎｅＢａｎｋ收录（ａｃｃｅｓｓｉｏｎＮｏＡＦ０４５０２３）。结论：该ＶＨ基因为抗人卵巢癌单抗ＣＯＣ１８３Ｂ２功能性重链可变区基因。

Objective: To amplify and sequence the heavy chain variable region (VH ) gene of monoclonal antibody (McAb) COC183B2 against human ovarian carcinoma. Methods: The VH gene of mouse McAb COC183B2 against human ovarian carcinoma was amplified by RTPCR . The PCR product was then cloned into PUC19 vector. The recombinants were sequenced by Sanger's method. The VH gene was compared with Gene Bank and published mouse VH genes. Results: It was proved that a fulllength VH gene was 354 bp, the VH gene was a member of mouse Ig heavy chain subgroup miscellaneous and originated from rearrangement of VH、 Dsp2.5 and MUSJH4 gene. The VH gene sequence was registered by Gene Bank(accession No. AF045023). Conclusion:It was suggested that obtained VH gene was potentially functional gene of the McAb COC183B2 against human ovarian carcinoma