Premium
The trp1 ‐ Δ FA designer deletion for PCR‐based gene functional analysis in Saccharomyces cerevisiae
Author(s) -
Horecka Joe,
Jigami Yoshifumi
Publication year - 1999
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/(sici)1097-0061(199912)15:16<1769::aid-yea495>3.0.co;2-m
Subject(s) - biology , gene , saccharomyces cerevisiae , genetics , homology (biology) , heterologous , genome , allele , computational biology , heterologous expression , microbiology and biotechnology , recombinant dna
PCR‐based gene deletion and modification are now common techniques for rapid gene manipulation in the yeast Saccharomyces cerevisiae . The techniques work best when the host strain lacks sequence homology to the PCR‐amplified selectable markers. One of the most versatile sets of PCR deletion/modification vectors is the pFA system described by Longtine et al. (1998), which is based on both heterologous ( kanMX6 and HIS3MX6 ) and homologous ( TRP1 ) markers. Here we describe the trp1‐ Δ FA designer deletion allele that removes precisely from the genome TRP1 sequences carried in the pFA vectors. The trp1‐ Δ FA allele can be introduced easily into TRP1 and most trp1 starting strains, and its use increases the frequency of correct integrants when using the pFA system's TRP1 ‐based constructs. Unlike trp1‐ Δ 1 , trp1 ‐Δ FA does not remove neighbouring GAL3 upstream activating sequences and therefore does not interfere with GAL gene induction. Copyright © 1999 John Wiley & Sons, Ltd.