title: Extraction of Chromosomal DNA from Schizosaccharomyces pombe.
abstract: Extraction of DNA from Schizosaccharomyces pombe cells is required for various uses, including templating polymerase chain reactions (PCRs), Southern blotting, library construction, and high-throughput sequencing. To purify high-quality DNA, the cell wall is removed by digestion with Zymolyase or Lyticase and the resulting spheroplasts lysed using sodium dodecyl sulfate (SDS). Cell debris, SDS, and SDS-protein complexes are subsequently precipitated by the addition of potassium acetate and removed by centrifugation. Finally, DNA is precipitated using isopropanol. At this stage, purity is usually sufficient for PCR. However, for more sensitive procedures, such as restriction enzyme digestion, additional purification steps, including proteinase K digestion and phenol-chloroform extraction, are recommended. All of these steps are described in detail here.
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Falmer, Brighton, E. Sussex BN1 9RQ, United Kingdom.
authors.forename: Johanne M
authors.initials: JM
authors.lastname: Murray
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Falmer, Brighton, E. Sussex BN1 9RQ, United Kingdom.
authors.forename: Adam T
authors.initials: AT
authors.lastname: Watson
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Falmer, Brighton, E. Sussex BN1 9RQ, United Kingdom.
authors.forename: Antony M
authors.initials: AM
authors.lastname: Carr
mesh_terms: D004271:DNA, Fungal / Q000302:isolation & purification*; D005827:Genetics, Microbial / Q000379:methods*; D008967:Molecular Biology / Q000379:methods*; D012568:Schizosaccharomyces / Q000235:genetics*
chemical_list: D004271:DNA, Fungal