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A family of vectors that facilitate transposon and insertional mutagenesis of cloned genes in yeast
Author(s) -
Allen James B.,
Elledge Stephen J.
Publication year - 1994
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/yea.320101003
Subject(s) - biology , ura3 , insertional mutagenesis , genetics , transposable element , selectable marker , plasmid , transposon mutagenesis , sleeping beauty transposon system , multiple cloning site , pbr322 , mutagenesis , gene , restriction site , mutation , vector (molecular biology) , restriction enzyme , mutant , recombinant dna
This report describes two sets of plasmid vectors that facilitate the identification of regions of complementation in cloned genomic inserts via transposon or insertional mutagenesis. The first set contains ARS ‐ H4 CEN6 , a yeast selectable nutritional marker ( HIS3, TRP1 or URA3 ), and neo for selection in Escherichia coli . These plasmids lack the Tn3 transposition immunity region present in pBR322 derived vectors, and are permissive recipients for Tn3 transposon mutagenesis. The second family of plasmids described facilitate gene disruption procedures performed in vitro . These vectors carry disruption cassettes that contain different yeast selectable markers ( HIS3, LEU2, TRP1 or URA3 ) adjacent to the Tn5 neo gene. These genes can be excised as a cassette on a common restriction fragment and introduced into any desired restriction site with selection for kanamycin resistance.