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Schizosaccharomyces
About: Schizosaccharomyces is a research topic. Over the lifetime, 2254 publications have been published within this topic receiving 123907 citations.
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TL;DR: This chapter describes techniques concerned with classical and molecular genetics, cell biology, and biochemistry that can be used with Schizosaccharomyces pombe.
Abstract: Publisher Summary This chapter describes techniques concerned with classical and molecular genetics, cell biology, and biochemistry that can be used with Schizosaccharomyces pombe . Conjugation and sporulation cannot take place in S. pombe except under conditions of nutrient starvation. ME medium is generally used for genetic crosses. To cross two strains, a loopful of h – and a loopful of h + are mixed together on a ME plate. The cross is left to dry and is then incubated below 30°, as conjugation is severely reduced above this temperature. Fully formed four-spore asci can be seen after 2–3 days of incubation. A 2 day-old cross is usually used for tetrad analysis. Using a 3-day-old cross, one can check for the presence of asci under the light microscope. Random spore analysis allows many more spores to be examined than in tetrad analysis, and in this way recombination mapping and strain construction can be carried out. Diploid cells arise spontaneously in most S. pombe strains, this characteristic can be used to isolate homozygous diploids of any strain. For mutagenesis of yeast strains, ethylmethane sulfonate (EMS) and nitrosoguanidine is used.
3,465 citations
TL;DR: A straightforward PCR‐based approach to the deletion, tagging, and overexpression of genes in their normal chromosomal locations in the fission yeast Schizosaccharomyces pombe, and a series of plasmids containing the kanMX6 module, which allows selection of G418‐resistant cells and thus provides a new heterologous marker for use in S. pom be.
Abstract: We describe a straightforward PCR-based approach to the deletion, tagging, and overexpression of genes in their normal chromosomal locations in the fission yeast Schizosaccharomyces pombe. Using this approach and the S. pombe ura4+ gene as a marker, nine genes were deleted with efficiencies of homologous integration ranging from 6 to 63%. We also constructed a series of plasmids containing the kanMX6 module, which allows selection of G418-resistant cells and thus provides a new heterologous marker for use in S. pombe. The modular nature of these constructs allows a small number of PCR primers to be used for a wide variety of gene manipulations, including deletion, overexpression (using the regulatable nmt1 promoter), C- or N-terminal protein tagging (with HA, Myc, GST, or GFP), and partial C- or N-terminal deletions with or without tagging. Nine genes were manipulated using these kanMX6 constructs as templates for PCR. The PCR primers included 60 to 80 bp of flanking sequences homologous to target sequences in the genome. Transformants were screened for homologous integration by PCR. In most cases, the efficiency of homologous integration was > or = 50%, and the lowest efficiency encountered was 17%. The methodology and constructs described here should greatly facilitate analysis of gene function in S. pombe.
2,212 citations
Wellcome Trust Sanger Institute1, London Research Institute2, Katholieke Universiteit Leuven3, Max Planck Society4, GATC Biotech5, Université catholique de Louvain6, Centre national de la recherche scientifique7, University of Exeter8, Institut national agronomique Paris Grignon9, Pablo de Olavide University10, University of Málaga11, University of Salamanca12, University of Sussex13, Salk Institute for Biological Studies14, Stanford University15, Cold Spring Harbor Laboratory16, TigerLogic17, Rosalind Franklin University of Medicine and Science18, Russian Academy of Sciences19, Technical University of Denmark20
TL;DR: The genome of fission yeast (Schizosaccharomyces pombe), which contains the smallest number of protein-coding genes yet recorded for a eukaryote, is sequenced and highly conserved genes important for eukARYotic cell organization including those required for the cytoskeleton, compartmentation, cell-cycle control, proteolysis, protein phosphorylation and RNA splicing are identified.
Abstract: We have sequenced and annotated the genome of fission yeast (Schizosaccharomyces pombe), which contains the smallest number of protein-coding genes yet recorded for a eukaryote: 4,824. The centromeres are between 35 and 110 kilobases (kb) and contain related repeats including a highly conserved 1.8-kb element. Regions upstream of genes are longer than in budding yeast (Saccharomyces cerevisiae), possibly reflecting more-extended control regions. Some 43% of the genes contain introns, of which there are 4,730. Fifty genes have significant similarity with human disease genes; half of these are cancer related. We identify highly conserved genes important for eukaryotic cell organization including those required for the cytoskeleton, compartmentation, cell-cycle control, proteolysis, protein phosphorylation and RNA splicing. These genes may have originated with the appearance of eukaryotic life. Few similarly conserved genes that are important for multicellular organization were identified, suggesting that the transition from prokaryotes to eukaryotes required more new genes than did the transition from unicellular to multicellular organization.
1,686 citations
TL;DR: The purification of complexes participating in heterochromatin formation has allowed us to begin to analyse in detail the processes involved, and in the future this will help to understand how the RNAi machinery acts to induce the chromatin modifications which lead to heterochROMatin assembly in fission yeast.
Abstract: In the fission yeast Schizosaccharomyces pombe, the RNAi pathway plays an important role in the formation and maintenance of heterochromatin Heterochromatin, or silent chromatin, is an epigenetically inherited attribute of eukaryotic chromosomes which is required for gene regulation, chromosome segregation and maintenance of genome stability In S pombe, heterochromatin forms on related repetitive DNA sequences at specific loci These repetitive sequences, in concert with the RNAi machinery, are thought to attract several proteins including chromatin-modifying enzymes which act to promote heterochromatin formation The purification of complexes participating in heterochromatin formation has allowed us to begin to analyse in detail the processes involved In the future this will help us to understand how the RNAi machinery acts to induce the chromatin modifications which lead to heterochromatin assembly in fission yeast
1,173 citations
TL;DR: The promoter and polyadenylation signal of the thiamine-repressible gene nmt1 of Schizosaccharomyces pombe have been used to construct the pREP extrachromosomally replicating plasmid and the pRIP integrative expression plasmids.
998 citations