Simple and efficient custom transcription activator-like effector gene synthesis via twin primer assembly.

Affiliation

Wang S(1), Yu Y(2), Pan Y(1), Wang H(1), Zhao Y(1), Qu M(1), Zhu Y(1).
Author information:
(1)Experimental & Translational Research Center, Beijing Friendship Hospital, Capital Medical University, Beijing 100050, China.
(2)Institute of Rehabilitation Science of China, China Rehabilitation Research Center, Beijing Key Laboratory of Neural Injury & Rehabilitation, Beijing 100068, China.

Abstract

Transcription activator-like effector (TALE) nucleases (TALENs) efficiently recognize and cleave DNA in a sequence-dependent manner. However, current TALE custom synthesis methods are either complicated or expensive. Here we report a simple and low-cost method for TALE construct assembly. This method utilizes the denaturation/reannealing nature of double-stranded DNA to create a unique single-stranded DNA overhang for proper ordering of TALE monomers in an engineered multimer. We successfully synthesized two TALEN pairs targeting the endogenous TET1 locus in human embryonic kidney cells and demonstrated their editing efficiency. Our method provides an alternative simple, low-cost method for effective TALEN assembly, which may improve the application of TALE-based technology.