DNA cloning by homologous recombination in Escherichia coli

Y. Zhang, J. P P Muyrers, G. Testa, A. F. Stewart

Research output: Contribution to journalArticle

Abstract

The cloning of foreign DNA in Escherichia coli episomes is a cornerstone of molecular biology. The pioneering work in the early 1970s, using DNA ligases to paste DNA into episomal vectors, is still the most widely used approach. Here we describe a different principle, using ET recombination, for directed cloning and subcloning, which offers a variety of advantages. Most prominently, a chosen DNA region can be cloned from a complex mixture without prior isolation. Hence cloning by ET recombination resembles PCR in that both involve the amplification of a DNA region between two chosen points. We apply the strategy to subclone chosen DNA regions from several target molecules resident in E. coli hosts, and to clone chosen DNA regions from genomic DNA preparations. Here we analyze basic aspects of the approach and present several examples that illustrate its simplicity, flexibility, and remarkable efficiency.

Original languageEnglish
Pages (from-to)1314-1317
Number of pages4
JournalNature Biotechnology
Volume18
Issue number12
DOIs
Publication statusPublished - 2000

Fingerprint

Cloning
Homologous Recombination
Escherichia coli
Organism Cloning
DNA
Genetic Recombination
DNA Ligases
Molecular biology
Ointments
Complex Mixtures
Amplification
Molecular Biology
Plasmids
Clone Cells
Polymerase Chain Reaction
Molecules

ASJC Scopus subject areas

  • Microbiology

Cite this

DNA cloning by homologous recombination in Escherichia coli. / Zhang, Y.; Muyrers, J. P P; Testa, G.; Stewart, A. F.

In: Nature Biotechnology, Vol. 18, No. 12, 2000, p. 1314-1317.

Research output: Contribution to journalArticle

Zhang, Y, Muyrers, JPP, Testa, G & Stewart, AF 2000, 'DNA cloning by homologous recombination in Escherichia coli', Nature Biotechnology, vol. 18, no. 12, pp. 1314-1317. https://doi.org/10.1038/82449
Zhang, Y. ; Muyrers, J. P P ; Testa, G. ; Stewart, A. F. / DNA cloning by homologous recombination in Escherichia coli. In: Nature Biotechnology. 2000 ; Vol. 18, No. 12. pp. 1314-1317.
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