Molecular characterization of a Saccharomyces plasmid containing theHIS4 gene

Self Cite

DNA was isolated from haploid strains of Saccharomyces cerevisiae carrying three different deletions in the his4 gene, his4-15, his4-24 and his4-29, as well as a his4 point mutant and a strain carrying the HIS4 wild type gene. Samples of the DNA preparations were treated with restriction endonucleases BamHl, EcoRl, Hindlll, PstI and SalI and the DNA fragments were separated according to size by agarose gel electrophoresis. A bacterial plasmid containing the yeast DNA sequences of a region including the HIS4 gene allowed the detection by molecular hybridization of the fragments in the electropherograms that contained these sequences. From the sizes of these fragments the mutations his4-15 and his4-24 were both determined to be deletions of about 0.5 kb, while his4-29 was found to be a deletion of approximately 0.9 kb. The data allowed an extension of our previously constructed cleavage map of the region as well as an approximate location of the deletions on the map.Comparisons with available data on the his4 gene and its product allowed an approximate positioning of the three functional regions of the his4 gene on the map and indicated that no large intervening sequences are present within the his4AB region.

Section: Resultsmentioning

confidence: 79%

“…In the accompanying paper (5) we have reported the construction of several plasmids which contain a 9.4 kb yeast chromosomal DNA sequence including the HIS4 gene. In one of these plasmids, pC502, the 9.4 kb segment is carried on the bacterial plasmid pBR325.…”

Section: Introductionmentioning

confidence: 99%

Molecular characterization of threehis4 deletion mutants in Saccharomyces cerevisiae

Holmberg

Kielland‐Brandt

Nilsson-Tillgren

et al. 1979

Self Cite

“…DNA fragments were transferred to nitrocellulose (Millipore HAWP 00010 or to Schleicher and Schuell BA85) by Southern transfer in 20 • SSC (33). The probes used in this study are pC503 (HIS4) ( 11,13) and the 0.9 kb EcoRI-SalI fragment (39) of pC512 (LEU2) (1,2,7,10,15,34). Both genes originate from chromosome III in S. cerevisiae.…”

Section: Molecular Hybridization Analysismentioning

confidence: 99%

DNA sequence polymorphisms in the genus Saccharomyces IV. Homoeologous chromosomes III of Saccharomyces bayanus, S. carlsbergensis, and S. uvarum

Pedersen

1986

Single chromosome III transfers have been accomplished from the two species S. bayanus and S. uvarum into genetically marked S. cerevisiae strains. Incompabilities between S. bayanus or S. uvarnm, and S. cerevisiae strains may account for the very few chromosome addition lines obtained. The preliminary genetic map for S. bayanus chromosome III is similar to the genetic map for a S. carlsbergensis chromosome lII from Danish lager strain BK2208. Recombination is absent between the transferred chromosomes and the auxotrophically marked chromosome from S. cerevisiae in the interval HIS4 to LEU2 but recombination does occur in the interval LEU2 to THR4. The mapped S. bayanus chromosome III carries HIS4 pattern III while the mapped chromosome III from the lager strain contains HIS4 pattern II. Transfer of a chromosome III from S. uvarum to S. cerevisiae has been recognised by restriction endonuclease fragment patterns, OFAGE chromosome separations and molecular hybridization, but a genetic map could not be constructed for an S. uvarum chromosome due to instability of the chromosome addition lines. The hypothesis is put forward that the S. carlsbergensis type strain and the lager strains are sibling species and species hybrids both produced by hybridization of an S. cerevisiae top fermenting strain and the bottom fermenting strain S. monacensis. The lager strains and the authentic type strain are designated S. carlsbergensis. From the electrophoretic karyotypes of type strains of Saecharomyces and molecular hybridization patterns it is deduced that S. bayanus, S. inusitatus, S. pastorianus and S. uvarum are closely related but distinct from the bottom fermenting strains. I N T R O D U C T I O NBase composition, DNA-RNA homology, DNA-DNA reassociation kinetics and restriction endonuclease fragment length polymorphisms have proven useful in the taxonomic evaluations of genera and species as well as in the diagnosis of closely related yeast strains (3,4,15,21,23,24,25,26,27,28,30,40). For taxonomic as well as breeding purposes the establishment of genetic linkage groups and their assignment to Abbreviations: OFAGE = Orthogonal field alternation gel electrophoresis; TBE = 0.089 M-Tris-borate, 0.089 M-boric acid, 0.002 M-EDTA; SSC = 0.15 M-NaC1, 15 mM-Na citrate.Springer-Verlag

“…For molecular hybridizations the plasmids pC503 containing HIS4 (22), pYF91.2 with LEU2 (45), and M271 with MATcz, (21) were used.…”

Section: Strains Recombinant Molecules Mediamentioning

confidence: 99%

“…(36). After gel electrophoresis, restriction endonuclease fragments ofgenomic yeast DNA were transferred to nitrocellulose filters (Millipore, HAWP 000 10) by the procedure of SOUTHERN (44) modified according to HOLMBERG et al (22). Hybridization of labelled DNA to DNA immobilized on nitrocellulose filters was performed as described by DENHARDT (8) with the modification of JEFFREYS and FLAVELL (25).…”

Section: In Vitro Labelling Of Dna Southern Transfers and Hybridizationmentioning

confidence: 99%

Genetic and molecular characterization of a distiller’s yeast

Keiding

1985

Auxotrophic mutants have been induced by UV-irradiation and EMS treatment in meiotic segregants of a distiller's production strain of yeast, In complementation tests some of the mutants were allelic to adel, ade3, lys2, met13, trpl, ural and ura4 of standard strains orS. cerevisiae.Crosses were made between two spore-derived clones of the distiller's strain and haploid S. cerevisiae strains carrying several auxotrophic markers. Tetrad analysis of the mating products revealed that the spore-derived clones were disomic for six and seven chromosomes, respectively.Chromosomes III from the distiller's production strain were compared to their counterparts in standard strains ofS. cerevisiae at several loci. Cloned DNA from HIS4, LEU2 and MATloci from the standard strain were used as hybridization probes to test the nucleotide sequence homology at these loci and the silent mating type genes HAIL and HMR. Restriction site polymorphisms were not detected inside the loci but in regions adjacent to them. These could be exploited in combination with tetrad analysis to identify four different chromosomes III in the distiller's strain. Of the four chromosomes, three were lacking HML and one was lacking HMR. The data are consistent with the assumption that all loci tested are organized in the same order as in the genetically analysed standard strains.