A counterselection for the tryptophan pathway in yeast: 5-fluoroanthranilic acid resistance.
The ability to counterselect, as well as to select for, a genetic marker has numerous applications in microbial genetics. Described here is the use of 5-fluoroanthranilic acid for the counterselection of TRP1, a commonly used genetic marker in the yeast Saccharomyces cerevisiae. Counterselection using 5-fluoroanthranilic acid involves antimetabolism by the enzymes of the tryptophan biosynthetic pathway, such that trp1, trp3, trp4 or trp5 strains, which lack enzymes required for the conversion of anthranilic acid to tryptophan, are resistant to 5-fluoroanthranilic acid. Commonly used genetic procedures, such as selection for loss of a chromosomally integrated plasmid, and a replica-plating method to rapidly assess genetic linkage in self-replicating shuttle vectors, can now be carried out using the TRP1 marker gene. In addition, novel tryptophan auxotrophs can be selected using 5-fluoroanthranilic acid.
Toyn
J H
JH
Department of Applied Biotechnology, DuPont Pharmaceuticals Co., Wilmington, DE 19880, USA. jeremy.h.toyn@dupontpharma.com
Gunyuzlu
P L
PL
White
W H
WH
Thompson
L A
LA
Hollis
G F
GF
D019747:Aldose-Ketose Isomerases; D000877:Anthranilate Phosphoribosyltransferase / Q000235:genetics; D000878:Anthranilate Synthase / Q000235:genetics; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D004352:Drug Resistance, Microbial / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics*; D017353:Gene Deletion; D005819:Genetic Markers; D007209:Indole-3-Glycerol-Phosphate Synthase / Q000235:genetics; D009154:Mutation; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D014364:Tryptophan / Q000096:biosynthesis*; D014367:Tryptophan Synthase / Q000235:genetics; D062367:ortho-Aminobenzoates / Q000737:chemistry / Q000494:pharmacology
D005656:Fungal Proteins; D005819:Genetic Markers; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D062367:ortho-Aminobenzoates; D014364:Tryptophan; D000877:Anthranilate Phosphoribosyltransferase; D007209:Indole-3-Glycerol-Phosphate Synthase; D000878:Anthranilate Synthase; D014367:Tryptophan Synthase; D019747:Aldose-Ketose Isomerases; C490353:TRP1 protein, S cerevisiae