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Updated: Jul 2, 2026
Construction of Synthetic Phage Displayed Fab Library with Tailored Diversity
Published on: May 1, 2018
Construction of a fully synthetic human scFv antibody library with CDR3 regions randomized by a split-mix-split
Chang-Cheng Yin1, Li-Li Ren, Lin-Lin Zhu
1National Laboratory of Biomacromolecules, and CAS-UT joint laboratory of Structural Virology and Immunology, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.
Abstract:
The randomization scheme of hypervariable region takes crucial role in construction of a synthetic antibody library. The codon bias and inevitable 'stop' codon of conventional 'NNK' and 'NNS' codons limit their applications. Here we report a split-mix-split DNA synthesis method that can control over the amino acid composition and distribution of randomized sequences effectually. A fully synthetic human antibody library with a diversity of 1.56 x 10(9) was successfully generated with complementarity determining region 3 (CDR3) randomized by this strategy. Sequencing analysis indicated that >60% of colonies had completely correct scFv genes and the amino acid composition and distribution were designed well in accordance. The utility was demonstrated by screening of scFv clones against BHL (anti-CD3 x anti-ovarian carcinoma bispecific antibody). These results proved the feasibility of the split-mix-split DNA randomization strategy in library construction and site-directed mutagenesis.
