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Verfasser Titel Jahr
zeige Details Maftahi, M.., Gaillardin, C.., ... Sticky-end polymerase chain reaction method for systematic gene disruption in Saccharomyces cerevisiae
in: Yeast, in: Yeast . - New York, NY [u.a.] : Wiley, ISSN 0749-503X, ZDB-ID 1479172-9 Vol. 12 (9. 1996), p. 859-868
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1996
We describe a new procedure for the generation of plasmids containing a large promoter and terminator region of a gene of interest, useful for gene disruption. In a two-step polymerase chain reaction (PCR), a fragment, corresponding to the terminator and promoter regions separated by a 16 bp sequence containing a rare restriction site (e.g. AscI), is synthesized (T-P fragment). This PCR fragment is cloned in vectors presenting a rare blunt-end cloning site and a yeast marker for selection in Saccharomyces cerevisiae (TRP1, HIS3 and KanMX). The final plasmids are used directly for gene disruption after linearization by the enzyme (e.g. AscI) specific for the rare restriction site. This approach was used to disrupt three open reading frames identified during the sequencing of COS14-1 from chromosome XIV of S. cerevisiae.
Copyright: Copyright © 1996 John Wiley & Sons Ltd.
beteiligte Personen: Maftahi, M.. , Gaillardin, C.. , Nicaud, J.-M..
Format: Elektronisch
Sprache: English
Erschienen: 1996.
Serie: Wiley InterScience Backfile Collection 1832-2000 [Dig. Serial]
Schlagwörter:
URL: http://dx.doi.org/10.1002/(SICI)1097-0061(199607)12:9<859::AID-YEA978>3.0.CO;2-Q

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