Abstract:

Abstract:【OBJECTIVE】To amplify various products of Fab antibody genes with diversity, guarantee enough reservoir capacity of phage antibody library.【METHOD】The spleen of C57BL\6 mouse was isolated, spleen cell suspension was prepared, total RNA was extracted, cDNA synthesis was accomplished by reverse transcription. The product of Fab antibody was amplified by PCR and Touch down PCR, the products were analyzed by agarose gel electrophoresis, we performed a comparative analysis of two PCR methods. 【RESULTS】the products of light chain and high chain gene was about 680bp, and the result corresponds with the theoretical value. The results of electrophoresis analysis showed that TD-PCR had 8 high chain gene and 6 light chain gene, PCR had 7 high chain gene and 5 light chain gene. The amplified gene product with VkB primer was not appeared in the products of PCR, and appeared in the products of TD-PCR. The gene product with IIIA and IIIC primer were obtained by TD-PCR, and not successfully amplified by PCR. The amplified gene product with IIIB primer was appeared in the products of PCR, and not appeared in the products of TD-PCR. 【 CONCLUSION 】TD-PCR may increase diversity of Fab antibody genes, is an approach to ensure reservoir capacity of phage antibody library，which would be a reference for amplification and other research of Fab antibody gene.