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[Protocol] Extraction of chromosomal DNA from Schizosaccharomyces pombe

journal contribution
posted on 2023-06-09, 09:19 authored by Jo Murray, Adam WatsonAdam Watson, Antony CarrAntony Carr
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.

Funding

Replication stalling in a palindrome; G0181; ASSOCIATION FOR INTERNATIONAL CANCER RESEARCH; 10-0273

Smc5/6 and replication fork stability; G0179; MRC-MEDICAL RESEARCH COUNCIL; G0901011

History

Publication status

  • Published

File Version

  • Accepted version

Journal

Cold Spring Harbor Protocols

ISSN

1559-6095

Publisher

Cold Spring Harbor Laboratory Press

Issue

5

Volume

2016

Department affiliated with

  • Sussex Centre for Genome Damage Stability Publications

Research groups affiliated with

  • Genome Damage and Stability Centre Publications

Full text available

  • No

Peer reviewed?

  • No

Legacy Posted Date

2017-12-13

First Compliant Deposit (FCD) Date

2017-12-13

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