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USER friendly DNA recombination (USERec): a simple and flexible near homology-independent method for gene library construction.

Villiers, B. R. M. ; Stein, Viktor ; Hollfelder, F. (2010)
USER friendly DNA recombination (USERec): a simple and flexible near homology-independent method for gene library construction.
In: Protein engineering, design & selection : PEDS, 23 (1)
Artikel, Bibliographie

Kurzbeschreibung (Abstract)

USER friendly DNA recombination (USERec) is introduced as a near homology-independent method that allows the simultaneous recombination of an unprecedented number of 10 DNA fragments (approximately 40-400 bp) within a day. The large number of fragments and their ease of preparation enables the creation of libraries of much larger genetic diversity (potentially approximately 10(10)-10(11) sequences) than current alternative methods based on DNA truncation (ITCHY, SCRATCHY and SHIPREC) or type IIb restriction enzymes (SISDC). At the same time, the frequency of frameshifts in the recombined library is low (90% of the recombined sequences are in frame). Compared to overlap extension PCR, USERec also requires much reduced crossover sequence constraints (only a 5'-AN(4-8)T-3' motif) and fewer experimental steps. Based on its simplicity and flexibility, and the accessibility of large and high quality recombined DNA libraries, USERec is established as a convenient alternative for the combinatorial assembly of gene fragments (e.g. exon or domain shuffling) and for a number of applications in gene library construction, such as loop grafting and multi-site-directed or random mutagenesis.

Typ des Eintrags: Artikel
Erschienen: 2010
Autor(en): Villiers, B. R. M. ; Stein, Viktor ; Hollfelder, F.
Art des Eintrags: Bibliographie
Titel: USER friendly DNA recombination (USERec): a simple and flexible near homology-independent method for gene library construction.
Sprache: Englisch
Publikationsjahr: 2010
Titel der Zeitschrift, Zeitung oder Schriftenreihe: Protein engineering, design & selection : PEDS
Jahrgang/Volume einer Zeitschrift: 23
(Heft-)Nummer: 1
Kurzbeschreibung (Abstract):

USER friendly DNA recombination (USERec) is introduced as a near homology-independent method that allows the simultaneous recombination of an unprecedented number of 10 DNA fragments (approximately 40-400 bp) within a day. The large number of fragments and their ease of preparation enables the creation of libraries of much larger genetic diversity (potentially approximately 10(10)-10(11) sequences) than current alternative methods based on DNA truncation (ITCHY, SCRATCHY and SHIPREC) or type IIb restriction enzymes (SISDC). At the same time, the frequency of frameshifts in the recombined library is low (90% of the recombined sequences are in frame). Compared to overlap extension PCR, USERec also requires much reduced crossover sequence constraints (only a 5'-AN(4-8)T-3' motif) and fewer experimental steps. Based on its simplicity and flexibility, and the accessibility of large and high quality recombined DNA libraries, USERec is established as a convenient alternative for the combinatorial assembly of gene fragments (e.g. exon or domain shuffling) and for a number of applications in gene library construction, such as loop grafting and multi-site-directed or random mutagenesis.

Fachbereich(e)/-gebiet(e): 10 Fachbereich Biologie
10 Fachbereich Biologie > Protein Engineering of Ion Conducting Nanopores
Hinterlegungsdatum: 14 Nov 2016 13:20
Letzte Änderung: 14 Nov 2016 13:20
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