Oct 05, 2025

Public workspaceDNA Extraction

DOI
https://dx.doi.org/10.17504/protocols.io.kxygx41odl8j/v1
  • igemnthuth 1
  • 1iGEM_NTHU_Taiwan
  • iGEM NTHU
igemnthuth
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DOI: https://dx.doi.org/10.17504/protocols.io.kxygx41odl8j/v1
Protocol Citation: igemnthuth 2025. DNA Extraction. protocols.io https://dx.doi.org/10.17504/protocols.io.kxygx41odl8j/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: October 03, 2025
Last Modified: October 05, 2025
Protocol Integer ID: 228980
Keywords: dna extraction extract plasmid dna, dna extraction extract, dna extraction, plasmid, dna, extraction
Abstract
Extract plasmid DNA from antibiotic-selected E. coli.
Troubleshooting
Pellet 1–5 ml bacterial overnight culture by centrifugation at >8000 rpm (6800 x g) for 3
min at room temperature (15–25°C).

Resuspend pelleted bacterial cells in 250 μl Buffer P1 and transfer to a microcentrifuge
tube.
Add 250 μl Buffer P2 and mix thoroughly by inverting the tube 4–6 times until the
solution becomes clear. Do not allow the lysis reaction to proceed for
more than 5 min. If using LyseBlue reagent, the solution will turn blue.


Add 350 μl Buffer N3 and mix immediately and thoroughly by inverting the tube 4–6
times. If using LyseBlue reagent, the solution will turn colorless.
Centrifuge for 10 min at 13,000 rpm (~17,900 x g) in a table-top microcentrifuge.
Apply 800 μl supernatant from step 5 to the QIAprep 2.0 spin column by pipetting. For