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-	<a name="The way it's done"></a><h4><b>The way it's done</b></h3>	+	<a name="The way it's done"></a><h4><b>The way it's done</b></h3><br>
-	The overlapping DNA molecules	+	The 5´-T5 exonuclease removes the bases from the 5'-end of double strained DNA molecule, leaving a recess in the DNA and resulting in useful ssDNA overhangs. The T5 exonuclease are inactivated during the incubation at 50C, whereas Phusion polymerase and Taq ligase filled the gaps of the annealed sequence and sealed the nicks.
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-	We describe an isothermal, single-reaction method for assembling multiple overlapping DNA molecules by the concerted action of a 5¢ exonuclease, a DNA polymerase and a DNA ligase. First we recessed DNA fragments, yielding single-stranded DNA overhangs that specifically annealed, and then covalently joined them. This assembly method can be used to seamlessly construct synthetic and natural genes, genetic pathways and entire genomes, and could be a useful molecular engineering tool.	+

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Revision as of 21:12, 5 September 2011

• Home
• About us
  • The Team
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• The Project
  • Abstract
  • Introduction
  • The Plug 'n' Play
    • Assembly system
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  • Other assembly systems
  • Achievements
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• Results
  • Background
  • Characterization
  • Proof of concept
    • Mammalian Cells
    • Fungi
  • Data page
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  • Collaboration
• Safety
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• Notebook
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• Our Partners
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  • Acknowledgements

	1 Gateway system 2 Gibson Assembly 3 In fusion 4 Sandwich 5 3A	Gibson Assembly Gibson Assembly is an isothermal, single-reaction method for assembling multiple overlapping DNA molecules, which was developed by Daniel G. Gibson at the J. Craig Venter Institute in 2009. The assembly system employs 5´-T5 exonuclease, PhusionDNA polymerase, and Taq lig and can be used to assemble both ssDNA and dsDNA. This methode makes it possible to join DNA molecules there are as large as 583kb and clone joined products in ''E. coli'' up to 300kb. The way it's done The 5´-T5 exonuclease removes the bases from the 5'-end of double strained DNA molecule, leaving a recess in the DNA and resulting in useful ssDNA overhangs. The T5 exonuclease are inactivated during the incubation at 50C, whereas Phusion polymerase and Taq ligase filled the gaps of the annealed sequence and sealed the nicks.