Patent ID: 7863222
Filing Date: 2011-01-04
Classification: C12N,C12P,C40B

Abstract:
1. A method of producing a library of nucleic acids, comprising: a) ligating a first double stranded adaptor comprising a first and a second type IIs restriction enzyme recognition sites to blunt end fragments to produce first adaptor-ligated fragments; b) digesting said adaptor-ligated fragments with said first type IIs restriction enzyme to produce digested fragments comprising said first double stranded adaptor and said second type IIs restriction enzyme recognition site; c) ligating a first hairpin adaptor comprising restriction enzyme recognition sites to said digested fragments to produce second adaptor-ligated fragments; d) digesting said second adaptor-ligated fragments with said second type IIs restriction enzyme to produce a population of hairpin-containing digestion products; e) ligating a second hairpin adaptor to said population of hairpin products to produce a population of circular nucleic acid molecules; f) amplifying said population of circular nucleic acid molecules using rolling circle amplification to produce a population of linear double stranded nucleic acids; g) digesting said population of linear double stranded nucleic acids with restriction enzymes that recognizes said restriction enzyme recognition sites of said first hairpin adaptor to produce a population of clonable fragments comprising nucleic acids of the formula: wherein: S h) ligating said clonable fragments into a vector to produce a population of circular vectors; i) digesting said population of circular vectors with said second restriction enzyme to produce digested vectors; and j) intramolecularly religating said digested vectors to produce said library of nucleic acids vectors comprising a plurality of nucleic acid molecules of the formula: wherein S1 and S2 are as set forth above, and R is a six base recognition site for said second restriction endonuclease.