Efficient preparation of internally modified single-molecule constructs using nicking enzymes.

ABSTRACT: Investigations of enzymes involved in DNA metabolism have strongly benefited from the establishment of single molecule techniques. These experiments frequently require elaborate DNA substrates, which carry chemical labels or nucleic acid tertiary structures. Preparing such constructs often represents a technical challenge: long modified DNA molecules are usually produced via multi-step processes, involving low efficiency intermolecular ligations of several fragments. Here, we show how long stretches of DNA (>50 bp) can be modified using nicking enzymes to produce complex DNA constructs. Multiple different chemical and structural modifications can be placed internally along DNA, in a specific and precise manner. Furthermore, the nicks created can be resealed efficiently yielding intact mole

SUBMITTER: Luzzietti N 

PROVIDER: S-EPMC3035433 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature