Cloning and Sequence Analysis of Light Variable Region Gene of Anti-human Retinoblastoma Monoclonal Antibody  

作　　者：Xiufeng Zhong, Yongping Li, Shuqi Huang, Bo Ning Chunyan Zhang, Jianliang Zheng, Guanguang FengZhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou 510060, China 

出　　处：《眼科学报》2002年第3期185-189,共5页Eye Science

基　　金：Supported in part by Grants 39870801,39400144 from National Natural Sciences Foundation of P.R.China;by Grants 98011 from Natural Sciences Foundation of Guangdong Province and by Grants 98007 from 211 Project Foundation of Sun Yat-Sen University of Medic

摘　　要：Purpose: To clone the variable region gene of light chain of monoclonal antibody against human retinoblastoma and to analyze the characterization of its nucleotide sequence as well as amino acid sequence.Methods: Total RNA was extracted from 3C6 hybridoma cells secreting specific monoclonal antibody(McAb)against human retinoblastoma(RB), then transcripted reversely into cDNA with olig-dT primers.The variable region of the light chain (VL) gene fragments was amplified using polymeerase chain reaction(PCR) and further cloned into pGEM(R) -T Easy vector. Then, 3C6 VL cDNA was sequenced by Sanger's method.Homologous analysis was done by NCBI BLAST.Results: The complete nucleotide sequence of 3C6 VL cDNA consisted of 321 bp encoding 107 amino acid residues, containing four workframe regions(FRs)and three complementarity-determining regions (CDRs) as well as the typical structure of two cys residues. The sequence is most homological to a member of the Vk9 gene family, and its chain utilizes the Jkl gene segment.Conclusion: The light chain variable region gene of the McAb against human RB was amplified successfully , which belongs to the Vk9 gene family and utilizes Vk-Jk1 gene rearrangement. This study lays a good basis for constructing a recombinant antibody and for making a new targeted therapeutic agents against retinoblastoma.Purpose:To clone the vanabie region gene ot light chain ot monoclonal antibody against human retinoblastoma and to analyze the characterization of its nucleotide sequence as well as amino acid sequence. Methods : Total RNA was extracted from 3C6 hybridoma cells secreting specific monoclonal antibody (MeAb) against human retinoblastoma (RB), then transcripted reversely into cDNA with olig-dT primers. The variable region of the light chain (VL) gene fragments was amplified using polymerase chain reaction (PCR) and further cloned into pGEM -T Easy vector. Then, 3C6 VL cDNA was sequenced by Sanger's method. Homologous analysis was done by NCBI BLAST. Results-. The complete nucleotide sequence of 3C6 VL cDNA consisted of 321 bp encoding 107 amino acid residues, containing four workframe regions (FRs) and three complementarity-determining regions (CDRs) as well as the typical structure of two cys residues. The sequence is most homological to a member of the Vk9 gene family , and its chain utilizes the Jk1 gene segment. Conclusion: The light chain variable region gene of the McAb against human RB was amplified successfully , which belongs to the Vk9 gene family and utilizes Vk-Jk1 gene rearrangement. This study lays a good basis for constructing a recombinant antibody and for making a new targeted therapeutic agents against retinoblastoma. Eye Science 2002; 18: 185 - 189.

关 键 词：抗人成视网膜细胞瘤 单克隆抗体 轻锭 可变区基因 基因克隆 序列分析 

分 类 号：R739.7[医药卫生—肿瘤]