Methodology

Rapid single step subcloning procedure by combined action of type II and type IIs endonucleases with ligase

Tobias Fromme and Martin Klingenspor

Department of Animal Physiology, Faculty of Biology, Philipps-University, D-35043 Marburg, Germany

author email corresponding author email

Journal of Biological Engineering 2007, 1:7doi:10.1186/1754-1611-1-7

Published:	26 November 2007

Abstract

Background

The subcloning of a DNA fragment from an entry vector into a destination vector is a routinely performed task in molecular biology labs.

Results

We here present a novel benchtop procedure to achieve rapid recombination into any destination vector of choice with the sole requirement of an endonuclease recognition site. The method relies on a specifically designed entry vector and the combined action of type II and type IIs endonucleases with ligase. The formulation leads to accumulation of a single stable cloning product representing the desired insert carrying destination vector.

Conclusion

The described method provides a fast single step procedure for routine subcloning from an entry vector into a series of destination vectors with the same restriction enzyme recognition site.