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In vitro recombination method

  • US 9,534,251 B2
  • Filed: 03/30/2010
  • Issued: 01/03/2017
  • Est. Priority Date: 08/11/2005
  • Status: Active Grant
First Claim
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1. A kit for in vitro joining a plurality of dsDNA molecules, consisting essentially of in a single container,a mixture of the isolated proteins(i) a single stranded DNA binding protein (SSB) which accelerates nucleic acid annealing,(ii) a non strand-displacing DNA polymerase, and(iii) a ligase;

  • (iv) an isolated non-processive 5′

    exonuclease; and

    (v) glycerol in an amount effective as a preservative;

    wherein the ratios of activities of (i), (ii), (iii), and (iv) are effective to achieve in vitro joining of dsDNA molecules, andwherein the dsDNA molecules are joined via a region of sequence homology to form a DNA molecule in which a single copy of the region of sequence homology is retained.

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