Codon-precise, synthetic, antibody fragment libraries built using automated hexamer codon additions and validated through next generation sequencing

Frigotto, Laura, Smith, Matthew E., Brankin, Christopher, Sedani, Ashni, Cooper, Simon E., Kanwar, Nisha, Evans, Daniel, Svobodova, Stanislava, Baar, Claudia, Glanville, Jacob, Ullman, Christopher G. and Hine, Anna V. (2015). Codon-precise, synthetic, antibody fragment libraries built using automated hexamer codon additions and validated through next generation sequencing. Antibodies, 4 (2), pp. 88-102.

Abstract

We have previously described ProxiMAX, a technology that enables the fabrication of precise, combinatorial gene libraries via codon-by-codon saturation mutagenesis. ProxiMAX was originally performed using manual, enzymatic transfer of codons via blunt-end ligation. Here we present Colibra™: an automated, proprietary version of ProxiMAX used specifically for antibody library generation, in which double-codon hexamers are transferred during the saturation cycling process. The reduction in process complexity, resulting library quality and an unprecedented saturation of up to 24 contiguous codons are described. Utility of the method is demonstrated via fabrication of complementarity determining regions (CDR) in antibody fragment libraries and next generation sequencing (NGS) analysis of their quality and diversity.

Publication DOI: https://doi.org/10.3390/antib4020088
Divisions: Life & Health Sciences > Biosciences
Life & Health Sciences
Life & Health Sciences > Cellular and Molecular Biomedicine
Additional Information: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Uncontrolled Keywords: colibra,proxiMAX randomization,saturation mutagenesis,CDR,single domain antibody,camelid antibody,antibody engineering
Full Text Link: http://www.mdpi ... 073-4468/4/2/88
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Published Date: 2015-05-15
Authors: Frigotto, Laura
Smith, Matthew E.
Brankin, Christopher
Sedani, Ashni
Cooper, Simon E.
Kanwar, Nisha
Evans, Daniel
Svobodova, Stanislava
Baar, Claudia
Glanville, Jacob
Ullman, Christopher G.
Hine, Anna V. ( 0000-0003-4065-831X)

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License: Creative Commons Attribution


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