Production of a Chimeric Mouse-Fish Monoclonal Antibody by the CRISPR/Cas9 Technology

Abstract

The CRISPR/Cas9 system, a defense mechanism naturally occurring in prokaryotes, has been recently repurposed as an RNA-guided DNA targeting platform and widely used as a powerful tool for genome editing. Here we describe how to modify the carboxy-terminal region, called Fragment crystallizable (Fc) region, of a murine monoclonal antibody by replacing the heavy chain constant exons with those from a teleost fish antibody by the CRISPR/Cas9 system. We outline optimal conditions for knockout and knockin mechanisms to edit the Immunoglobulin heavy chain (IgH) constant region gene locus in a murine hybridoma cell line. A chimeric mouse-fish monoclonal antibody can be successfully produced by hybridoma cell lines engineered according to this protocol.

Keywords: CRISPR/Cas9; Fc region; Genome editing; IgH gene locus; Monoclonal antibody; Murine hybridoma; Teleost fish.