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Simultaneous Removal of Multiple DNA Segments by Polymerase Chain Reactions

Part of the Methods in Molecular Biology book series (MIMB,volume 1472)

Abstract

Precise DNA manipulation is a key enabling technology for synthetic biology. Approaches based on restriction digestion are often limited by the presence of certain restriction enzyme recognition sites. Recent development of restriction-free cloning approaches has greatly enhanced the flexibility and speed of molecular cloning. Most restriction-free cloning methods focus on DNA assembly. Much less work has been dedicated towards DNA removal. Here we introduce a protocol that allows simultaneous removal of multiple DNA segments from a plasmid using polymerase chain reactions (PCR). Our approach will be beneficial to applications in multiple sites mutagenesis, DNA library construction, genetic and protein engineering, and synthetic biology.

Key words

  • Restriction-free cloning
  • Polymerase chain reaction
  • Synthetic DNA assembly and manipulation
  • Multiplex gene removal
  • Synthetic single-stranded bridging oligos

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  • DOI: 10.1007/978-1-4939-6343-0_15
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Acknowledgements

K.Z. thanks the funding support from the University of Illinois at Urbana-Champaign (UIUC).

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Correspondence to Kai Zhang Ph.D. .

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Krishnamurthy, V., Zhang, K. (2017). Simultaneous Removal of Multiple DNA Segments by Polymerase Chain Reactions. In: Hughes, R. (eds) Synthetic DNA. Methods in Molecular Biology, vol 1472. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6343-0_15

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  • DOI: https://doi.org/10.1007/978-1-4939-6343-0_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6341-6

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