Simultaneous removal of multiple DNA segments by polymerase chain reactions

Vishnu Krishnamurthy, Kai Zhang

Research output: Chapter in Book/Report/Conference proceedingChapter

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.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages193-203
Number of pages11
DOIs
StatePublished - Jan 1 2017

Publication series

NameMethods in Molecular Biology
Volume1472
ISSN (Print)1064-3745

Fingerprint

DNA-Directed DNA Polymerase
Synthetic Biology
Polymerase Chain Reaction
DNA
Organism Cloning
Protein Engineering
Molecular Cloning
Gene Library
Mutagenesis
Digestion
Plasmids
Technology
Enzymes

Keywords

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

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Krishnamurthy, V., & Zhang, K. (2017). Simultaneous removal of multiple DNA segments by polymerase chain reactions. In Methods in Molecular Biology (pp. 193-203). (Methods in Molecular Biology; Vol. 1472). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-6343-0_15

Simultaneous removal of multiple DNA segments by polymerase chain reactions. / Krishnamurthy, Vishnu; Zhang, Kai.

Methods in Molecular Biology. Humana Press Inc., 2017. p. 193-203 (Methods in Molecular Biology; Vol. 1472).

Research output: Chapter in Book/Report/Conference proceedingChapter

Krishnamurthy, V & Zhang, K 2017, Simultaneous removal of multiple DNA segments by polymerase chain reactions. in Methods in Molecular Biology. Methods in Molecular Biology, vol. 1472, Humana Press Inc., pp. 193-203. https://doi.org/10.1007/978-1-4939-6343-0_15
Krishnamurthy V, Zhang K. Simultaneous removal of multiple DNA segments by polymerase chain reactions. In Methods in Molecular Biology. Humana Press Inc. 2017. p. 193-203. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-4939-6343-0_15
Krishnamurthy, Vishnu ; Zhang, Kai. / Simultaneous removal of multiple DNA segments by polymerase chain reactions. Methods in Molecular Biology. Humana Press Inc., 2017. pp. 193-203 (Methods in Molecular Biology).
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