[Cloning and sequencing of VH and VL genes of an anti-human lung adenocarcinoma monoclonal antibody].
By comparing the conserved regions at each end of the nucleotide sequences of murine germ-line genes encoding FR1 and FR4 regions of immunoglobulin heavy and light chain variable regions, we designed two sets of primer for amplification of VH and VL genes. Hybridoma cell WLA-2C4, secreting an antihuman lung adenocarcinoma McAb, was cultured and the genome DNA was extracted and used as template for PCR. After PCR the desired VH and VL fragments were amplified. The PCR products were then cloned into pUC19 vector. By screening and identification, several recombinants that had been inserted with the target fragments were obtained. Then they were sequenced with Sanger's method. It was confirmed by computer-assisted comparative sequence analysis that clones of full-length and potentially functional VH and VL genes from the hybridoma cell line WLA-2C4 were ob-tained.
Wang
Y
Y
Institute of Genetics, Academia Sinica, Beijing.
Liu
X
X
Gu
Z
Z
Xiao
S
S
Chen
A
A
Lin
Q
Q
Huang
H
H
Wang
D
D
Li
X
X
D000230:Adenocarcinoma / Q000276:immunology*; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D005803:Genes, Immunoglobulin; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D007147:Immunoglobulin Light Chains / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D008175:Lung Neoplasms / Q000276:immunology*; D051379:Mice; D008969:Molecular Sequence Data
D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region