METHODS AND COMPOSITIONS FOR RAPID SEAMLESS DNA ASSEMBLY

摘要

Provided herein are methods for assembling DNA fragments employing at least three enzymatic activities: DNA polymerase, flap endonuclease, and DNA ligase. Certain aspects include methods for generating closed circular DNA products, e.g., plasmid vectors, by assembling various DNA fragments having complementary ends that hybridize to one another. The resulting circular products can be introduced into host cells and selected for desired properties. Kits for performing the method are also provided.

主权项

1. A DNA assembly method comprising:
(a) obtaining at least two DNA fragments, wherein the at least two DNA fragments comprise ends that can selectively hybridize with one another; and (b) contacting the at least two DNA fragments with:
(i) a DNA polymerase;(ii) a flap endonuclease; and(iii) a DNA ligase;under reaction conditions that promote hybridization of the at least two DNA fragments and support the activities of components (i), (ii) and (iii), to produce a product comprising an assembled circular DNA comprising at least a portion of each of the at least two DNA fragments.