基础医学与临床2012，Vol.32Issue(1)：56-60,5.
引入易错PCR和DNA-shuffling技术构建单链抗体库
The introduction of error-prone PCR and DNA-shuffling technology to build mono-chain antibody library
杨浩 1杨青平 2查成喜 1韩跃武1
作者信息
- 1. 兰州大学 基础医学院
- 2. 甘肃省第三人民医院 内分泌科,甘肃 兰州 730000
- 折叠
摘要
Abstract
Objective To construct a mono-storage capacity single-chain antibody (scFv) library using error-prone PCR and DNA-shuffling technologies. Methods Peripheral blood was collected from different age, gender and healthy people. Total RNA was extracted from human peripheral blood mononuclear cells and reversely transcribed to cDNA by RT-PCR. VH and VL genes were amplified by PCR and mutated by error-prone PCR. The full-length ScFv fragments were recombinated with the DNA-shuffling technology in vitro, and then connected with T vector and transformed into E. Coli DH5α. Antibody library diversity was identified by restriction enzyme digestion after colony PCR, and then the capacity of scFv was calculated. Results The 4. 37 x 1013 capacity and better diversity ScFv library was constructed by error-prone PCR and DNA-shuffling techniques. Conclusions The scFv library with high-storage capacity and genetic diversity has been constructed, which laid a foundation of selection high-affinity antibodies.关键词
易错PCR/DNA-shuffling/单链抗体库Key words
error-prone PCR/DNA-shuffling/scFv library分类
医药卫生引用本文复制引用
杨浩,杨青平,查成喜,韩跃武..引入易错PCR和DNA-shuffling技术构建单链抗体库[J].基础医学与临床,2012,32(1):56-60,5.
