![]() Use lithium chloride (0.8 M final conc) for RNA.Use sodium chloride (0.2 M final conc) for DNA samples containing SDS, since NaCl keeps SDS soluble in 70% ethanol so that it doesn’t precipitate with the DNA.Use sodium acetate (0.3 M final conc, pH 5.2) for routine DNA precipitation.This step is to wash any residual salt away from the pelleted DNA. (Molecular Cloning, A Laboratory Manual 2 nd Edition… 2 nd edition? – I need to get a newer version!), this is not required, as nucleic acids at concentrations as low as 20 ng/mL will precipitate at 0–4☌, so incubation for 15–30 minutes on ice is sufficient. –20° or –80☌) is commonly cited as a necessary step in protocols. Incubation of the nucleic acid/salt/ethanol mixture at low temperatures (e.g. This shields its charge and makes the nucleic acid less hydrophilic, thus causing it to drop out of the solution. Ethanol, on the other hand, has a much lower dielectric constant, making it much easier for Na + to interact with the PO 4 –. Water has a high dielectric constant, which makes it fairly difficult for the Na + and PO 4 – to come together. The electrostatic attraction between the Na + ions in solution and the PO 4 – ions are dictated by Coulomb’s Law, which is affected by the dielectric constant of the solution. The positively charged sodium ions neutralize the negative charge on the PO 4 – groups on the nucleic acids, making the molecule far less hydrophilic and, therefore, much less soluble in water. In solution, sodium acetate breaks up into Na + and –. ![]() The role of salt in the protocol is to neutralize the charges on the sugar-phosphate backbone. Nucleic acids are hydrophilic due to the negatively charged phosphate (PO 4 –) groups along the sugar-phosphate backbone. Polar molecules can therefore be described as hydrophilic and non-polar molecules, which can’t easily interact with water molecules, are hydrophobic. Because of these charges, polar molecules like DNA or RNA can interact electrostatically with the water molecules, allowing them to easily dissolve in water. Water is a polar molecule – it has a partial negative charge near the oxygen atom due to the unshared pairs of electrons, and partial positive charges near the hydrogen atoms. So how does this work? It’s All About Solubility…įirst, we need to know why nucleic acids are soluble in water. After a further centrifugation step, the ethanol is removed and the nucleic acid pellet is allowed to dry before resuspending in a clean aqueous buffer. The pellet is then washed in cold 70% ethanol. After precipitation, the nucleic acids can then be separated from the rest of the solution by centrifugation. The basic procedure is that salt and ethanol are added to the aqueous solution, which forces the precipitation of nucleic acids out of the solution. What is Ethanol Precipitation of DNA?Įthanol precipitation is a commonly used technique for concentrating and de-salting nucleic acid (DNA or RNA) preparations in an aqueous solution. Ethanol precipitation of DNA and RNA works by adding salt and ethanol to a DNA solution, which makes the DNA less hydrophilic, causing it to precipitate out. DNA and RNA are hydrophilic (water-loving) molecules and, therefore, soluble in water.
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