Process of clean cloning
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United States of America Patent
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Nov 11, 2014
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app pub date -
Oct 8, 2009
filing date -
Oct 8, 2008
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Abstract
A process of inserting a nucleic acid sequence of interest into an acceptor nucleic acid is provided. The process comprises amplifying by PCR a DNA comprising in the following order a sequence segment U, a nucleic acid sequence segment of known nucleotide sequence K2, and a nucleic acid sequence segment of known sequence K3. The process further comprises treating the linear double-stranded DNA molecules from the PCR amplification with an exonuclease to obtain a single-stranded overhang at the first end of the DNA and a single-stranded overhang comprising nucleic acid segments K2 and K3 at the second end of the DNA. The process additionally comprises annealing the product of the exonuclease treatment to a linearized double-stranded acceptor nucleic acid which has been designed to complement the single-stranded overhangs of the product of the exonuclease treatment.
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