Rapid isolation of rare clones from highly complex DNA libraries by PCR analysis of liquid gel pools.
A semiliquid medium was employed in an efficient method to screen highly complex DNA libraries for clones with known sequences. Unbiased clone pools are generated in 2 ml vials and screened by whole-cell PCR, and individual clones are obtained by few additional rounds of dilution and PCR screening. To demonstrate the utility of this approach, the single positive clone present in a 400,000 member metagenomic fosmid library was isolated.
Hrvatin
Sinisa
S
Harvard College, Cambridge, USA.
Piel
Jörn
J
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002999:Clone Cells; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D015723:Gene Library; D016133:Polymerase Chain Reaction; D011161:Porifera / Q000235:genetics*
D004247:DNA