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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 CarrExtraction 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 ProtocolsISSN
1559-6095Publisher
Cold Spring Harbor Laboratory PressExternal DOI
Issue
5Volume
2016Department 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
2017-12-13First Compliant Deposit (FCD) Date
2017-12-13Usage metrics
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