[Amplification, cloning and sequence analysis of the variable region genes of monoclonal antibody against human bladder carcinoma].
We have designed two sets of oligonucleotide primers to amplify the immunoglobulin heavy- and light-chain variable region genes from genomic DNA by the polymerase chain reaction, PCR. The genomic DNA was extracted from hybridoma BDI-1 cell, which secrets a monoclonal antibody against human bladder carcinoma. The primers contain special restriction sites that allow the variable region genes to be easy to cloning for sequencing and expression. The PCR products were ligated to plasmid PUC19. The recombinants were sequenced with Sanger's method. It was proved that a full-length VH and V kappa genes was 366 and 324 base pairs respectively. Comparaing with other published sequences, the VH gene was a member of mouse heavy-chain VH subgroup II and originated from rearrangement of VH, Dsp2.2 and JH4; the V kappa gene was V kappa subgroup IV and from V kappa and J kappa 4. It was suggested that obtained VH and V kappa genes were potentially functional genes of the monoclonal antibody against human bladder carcinoma.
Xiao
S
S
Institute of Genetics, Academia Sinica, Beijing.
Yu
L
L
Gu
Z
Z
D000911:Antibodies, Monoclonal / Q000235:genetics*; D003001:Cloning, Molecular; D005784:Gene Amplification; D015321:Gene Rearrangement; D005803:Genes, Immunoglobulin; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000235:genetics; D007145:Immunoglobulin kappa-Chains / Q000235:genetics; D017422:Sequence Analysis, DNA; D014407:Tumor Cells, Cultured; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000276:immunology*
D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D007145:Immunoglobulin kappa-Chains