Feb 27, 2023

Public workspaceEthanol precipitation of nucleic acid

This protocol is a draft, published without a DOI.
  • 1University of Oslo
Andreas Sagen
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Protocol Citation: Andreas Sagen 2023. Ethanol precipitation of nucleic acid. protocols.io https://protocols.io/view/ethanol-precipitation-of-nucleic-acid-cn9evh3e
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: In development
We are still developing and optimizing this protocol
Created: February 10, 2023
Last Modified: February 27, 2023
Protocol Integer ID: 76806
Abstract
Simple protocol to precipitate nucleic acids
Protocol materials
ReagentGlycogenMerck MilliporeSigma (Sigma-Aldrich)Catalog #G1767
Step 6
ReagentSodium acetateMerck MilliporeSigma (Sigma-Aldrich)Catalog #S2889
In 2 steps
ReagentAcetic acidMerck MilliporeSigma (Sigma-Aldrich)Catalog #A6283
Step 3
Sodium acetate (3 M, pH 5.2)
Sodium acetate (3 M, pH 5.2)
Add Amount25 mL distilled water to a Amount50 mL canonical tube
Measure Amount12.305 g Sodium acetate

Materials:
ReagentSodium acetateSigma AldrichCatalog #S2889
Adjust pH to Ph5.2 with glacial acetic acid

Materials:
ReagentAcetic acidSigma AldrichCatalog #A6283
Autoclave solution at Temperature121 °C for Duration00:15:00
15m
Precipitation
Precipitation
2h
2h
Add Amount100 µL lysate in a reaction tube
Add Amount10 µL Sodium acetate (Concentration3 Molarity (M) , Ph5.2 ), Amount2 µL 250 µg/mL glycogen and Amount333 µL absolute ethanol

Material:
ReagentGlycogenMerck MilliporeSigma (Sigma-Aldrich)Catalog #G1767
ReagentSodium acetateMerck MilliporeSigma (Sigma-Aldrich)Catalog #S2889
Vortex solution, then precipitate at Temperature-20 °C or Temperature-80 °C for Duration01:00:00 or DurationOvernight

Note
Longer incubation and lower temperature provide better recovery of nucleic acids.

1h
Centrifuge at Centrifigation12000 rcf, 4°C, 00:30:00 . Discard supernatant, and repeat step 9.1 and 9.2, 2 times in total
30m
Discard supernatant, and resuspend pellet in Amount500 µL 70% absolute ethanol
Centrifuge at Centrifigation12000 rcf, 4°C, 00:30:00
30m
Air-dry the pellet

Note
Avoid over-drying the pellet, as it then takes more time to dissolve.

Resuspend pellet in Amount20 µL TE-buffer or nuclease-free water