Highly efficient preparation of single-stranded DNA rings by T4 ligase at abnormally low Mg(II) concentration.

ABSTRACT: Preparation of large amount of single-stranded circular DNA in high selectivity is crucial for further developments of nanotechnology and other DNA sciences. Herein, a simple but practically useful methodology to prepare DNA rings has been presented. One of the essential factors is to use highly diluted T4 ligase buffer for ligase reactions. This strategy is based on our unexpected finding that, in diluted T4 buffers, intermolecular polymerization of DNA fragments is greatly suppressed with respect to their intramolecular cyclization. This promotion of cyclization is attributable to abnormally low concentration of Mg2+ ion (0.5-1.0 mM) but not ATP in the media for T4 ligase reactions. The second essential factor is to add DNA substrate intermittently to the mixture and maintain its tempora

SUBMITTER: An R 

PROVIDER: S-EPMC5587803 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature