We report the synthesis of an ortho-phthalaldehyde (OPA) functionalized and water-soluble copolymer by reversible addition-fragmentation chain transfer (RAFT) copolymerization of a novel OPA derivative vinyl monomer with N,N-dimethylacrylamide, followed by deprotection in trifluoroacetic acid/dichloromethane solution. Our results demonstrate that the OPA-containing copolymer could conjugate rapidly, chemoselectively and efficiently with native proteins via the OPA-primary amine reaction under physiological conditions without significant changes in the protein activity. This work provides a promising method for efficient conjugation of polymers with native proteins under physiological conditions.