Versatile broad-host-range cosmids for construction of high quality metagenomic libraries

We constructed IncP broad-host-range Gateway (R) entry cosmids pJC8 and pJC24, which replicate in diverse Proteobacteria. We demonstrate the functionality of these vectors by extracting, purifying, and size-selecting metagenomic DNA from agricultural corn and wheat soils, followed by cloning into pJC8. Metagenomic DNA libraries of 8 x 10(4) (corn soil) and 9 x 10(6) (wheat soil) clones were generated for functional screening. The DNA cloned in these libraries can be transferred from these recombinant cosmids to Gateway destination vectors for specialized screening purposes. Those library clones are available from the Canadian MetaMicroBiome Library project (http://www.cm2bl.org/). (C) 2014 Elsevier B.V. All rights reserved.