Light and heavy chain expression vector backbone preparation

Catalent plasmid pRW449 (encoding human antibody #1 kappa light chain) was used as the starting material for generating the light chain vector backbone. pRW449 was digested with BamH1-HF (NEB) and DraIII-HF (NEB) to remove the wild type human kappa light chain constant region. The digested plasmid DNA was purified by 1% agarose gel electrophoresis and QIAquick gel extraction kit (QIAGEN). The purified DNA was used as a vector backbone for cloning the human IgG1 kappa light chain constant region containing the aldehyde tag at different positions.

Catalent plasmid pRW1064 (encoding human antibody #1 IgG1 heavy chain) was used as the starting material for the generation of the heavy chain vector backbone. pRW1064 was digested with KpnI-HF (NEB) and DraIII-HF to remove the wild-type human IgG1 heavy chain constant region. The digested plasmid DNA was purified by 1% agarose gel electrophoresis and QIAquick gel extraction kit. The purified DNA was used as a vector backbone for cloning human IgG1 heavy chain constant regions containing the aldehyde tag at different positions.