AFEAP cloning: a precise and efficient method for large DNA sequence assembly.

ABSTRACT:

Background

Recent development of DNA assembly technologies has spurred myriad advances in synthetic biology, but new tools are always required for complicated scenarios. Here, we have developed an alternative DNA assembly method named AFEAP cloning (Assembly of Fragment Ends After PCR), which allows scarless, modular, and reliable construction of biological pathways and circuits from basic genetic parts.

Methods

The AFEAP method requires two-round of PCRs followed by ligation of the sticky ends of DNA fragments. The first PCR yields linear DNA fragments and is followed by a second asymmetric (one primer) PCR and subsequent annealing that inserts overlapping overhangs at both sides of each DNA fragment. The overlapping overhangs of the neighboring DNA fragments annealed and t

SUBMITTER: Zeng F 

PROVIDER: S-EPMC5686892 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature