Highly efficient one-step scarless protein tagging by type IIS restriction endonuclease-mediated precision cloning.

ABSTRACT: Protein tagging with a wide variety of epitopes and/or fusion partners is used routinely to dissect protein function molecularly. Frequently, the required DNA subcloning is inefficient, especially in cases where multiple constructs are desired for a given protein with unique tags. Additionally, the generated clones have unwanted junction sequences introduced. To add versatile tags into the extracellular domain of the transmembrane protein THSD1, we developed a protein tagging technique that utilizes non-classical type IIS restriction enzymes that recognize non-palindromic DNA sequences and cleave outside of their recognition sites. Our results demonstrate that this method is highly efficient and can precisely fuse any tag into any position of a protein in a scarless manner. Moreover, this

SUBMITTER: Xu Z 

PROVIDER: S-EPMC5559197 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature