[Cloning and sequence characterization of heavy chain variable region gene from hybridoma cells secreting MAB against VEGF].
OBJECTIVE
To clone the variable region of heavy chain (VH) gene from hybridoma secreting monoclonal antibody against human vasoendothelial growth factor (VEGF) for construction of recombinant antibody (scFv).


METHODS
One step method was used to extract total RNA and degeneration primer was used to amplify the cDNAs by retropolymerase chain reaction (RT-PCR), then the products were inserted to T-vector and its sequence was analysed.


RESULTS
The VH gene contained 369 base pairs encoding 123 amine acid resides. Analysis of the nucleotide sequence of the light chain variable region revealed that the VH sequence was 87% identical to the sequence present in mouse Ig VH region.


CONCLUSIONS
The VH gene was 369 bp. According to Kabat classification, the VH gene belonging to mouse kappa light chains subgroup I (A), resulting from VH-D-JH4 rearrangment.
Yang
X
X
Department of Oral and Maxillofacial Surgery, College of Stomatology, West China University of Medical Sciences.
Zheng
G
G
Wang
D
D
Cheng
Y
Y
Li
B
B
Li
J
J
Fang
S
S
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics*; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D016228:Endothelial Growth Factors / Q000276:immunology*; D006801:Humans; D006825:Hybridomas / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D036341:Intercellular Signaling Peptides and Proteins / Q000276:immunology*; D008222:Lymphokines / Q000276:immunology*; D051379:Mice; D008969:Molecular Sequence Data; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D000911:Antibodies, Monoclonal; D016228:Endothelial Growth Factors; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D036341:Intercellular Signaling Peptides and Proteins; D008222:Lymphokines; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors