| 摘要 |
PROBLEM TO BE SOLVED: To provide a nucleic acid ligation method ligating two kinds of DNA or DNA library precisely and in high efficiency, and a screening method of a gene encoding a functional protein utilizing the ligation method, and a mass production method of the functional protein.SOLUTION: To a 5' side of a first DNA reverse primer and a 5' side of a second DNA forward primer, respectively, a linker sequence is provided and 5' terminal is phosphorylated. After amplifying each of the first and the second DNAs with PCR, the phosphorylated terminal chain is λ exonuclease-digested until the linker region becomes a single chain, annealed and substitution synthesized with BstDNA polymerase, and a double-stranded fusion DNA is produced. By selecting two kinds of DNA libraries as the first and the second DNAs, a double-stranded fusion DNA library is prepared, thereby a fusion protein library can be prepared by combining with a phage display method, so that the functional protein can be screened using target functionality as an index. Further by repeating the screening, the target functional protein can be selectively obtained. For example, a single chain antibody having high linkage activity to a target substance can be produced. |
