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Plasmid DNA Extraction from E. coli Using Alkaline Lysis Method
Published: Feb 5, 2011 DOI: 10.21769/BioProtoc.30 Views: 206917
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Abstract
This is a quick and efficient way to extract E. coli plasmid DNA without using commercial kits. This technique was invented by Birnboim and Doly (1979).
Materials and Reagents
- RNAase (Life Technologies, Invitrogen™)
- Isopropanol (EM Science)
- Ethanol Absolute (200 Proof) (VWR Chemical)
- Tryptone
- Yeast extract
- NaCl
- Glucose
- EDTA
- 0.2 N NaOH
- SDS
- KOAc
- Potassium acetate
- Glacial acetate
- Tris-HCl (pH 8.0)
- Luria-Bertani broth (LB) medium: Bacto-tryptone (BD Biosciences), yeast extract (BD Biosciences) (see Recipes)
- Resuspension solution (P1 buffer) (see Recipes)
- Lysis solution (P2 buffer) (see Recipes)
- Neutralizing solution (P3 buffer) (see Recipes)
- TE (see Recipes)
Equipment
- Table-top centrifuges
- 1.5-ml eppendorf tube
Note: Use the highest speed for all centrifugation steps in this protocol. - 37 °C heat blocker
Procedure
Copyright: © 2011 The Authors; exclusive licensee Bio-protocol LLC.
Category
Microbiology > Microbial genetics > DNA
Molecular Biology > DNA > DNA extraction
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