It is well known that Ethanol has a lower dielectric constant than water, making it to promote ionic bond formations between the Na+ (from the salt) and the PO3- (from the DNA backbone), further, causing the DNA to precipitate.Then, how the ethanol wash of the DNA pellet removes its salts at the end step of DNA isolation ? Is there any effect of reaction environment on ethanol that it promotes salt addition at sometime and removal at other ?
The initial role of the ethanol and monovalent cations is to remove the solvation shell surrounding the DNA and permitting the precipitation of the DNA in pellet form. The ethanol also serves to promote the aggregation of the DNA. With respect to the washing steps, typically a 70% ethanol solution is used. This permits the solubilisation of the salts whilst minimising the solubility of the DNA. The salts are therefore removed due to solubility differences, especially with the aggregated DNA. The final 100% ethanol wash that is usually employed serves more to permit the easier evaporation of the ethanol from the DNA pellet in order to prevent any carry over.
Actually, the ethanol rinse is intended to remove carryover salt from the precipitation step, not to dissociate sodium ions from backbone phosphates. The precipitated nucleic acids do remain in salt form.
Hope it helps!
before DNA elution, we let the eppendorf tube to dry until we see that there are no drops of ethanol after that we add elution buffer and that all.
Hope this proves helpful,
http://bitesizebio.com/2839/dna-precipitation-ethanol-vs-isopropanol/
Thanks everybody but i need more clarification about ...how ethanol removes salts ?and is it only NaCl or other salts which come with DNA?
The initial role of the ethanol and monovalent cations is to remove the solvation shell surrounding the DNA and permitting the precipitation of the DNA in pellet form. The ethanol also serves to promote the aggregation of the DNA. With respect to the washing steps, typically a 70% ethanol solution is used. This permits the solubilisation of the salts whilst minimising the solubility of the DNA. The salts are therefore removed due to solubility differences, especially with the aggregated DNA. The final 100% ethanol wash that is usually employed serves more to permit the easier evaporation of the ethanol from the DNA pellet in order to prevent any carry over.
in general organic solvents dissolve organic chemicals and ionic solvents dissolve ionic solutes so orgnic compounds like benzene dissolve in chloroform and sodium chloride does not while NaCl dissolves in water and benzene does not,
Ethanol has an organic CH3CH2 end and a partially charged OH moiety so it can dissolve /mix with water so the effect of washing precipitated DNA creates an environment where the charged dna stays insoluble due to the presence of the alcohol but the precipitated salts dissolve easily in the aqueous part of the mixture so can be easily removed by decantation
Re: how the wash works.
The pellet still contains traces of the solution used to precipitate the DNA, which contains trace amounts of NaCl (or whatever salt you used) that, in some cases, may inhibit downstream enzyme reactions (i.e. ammonium acetate inhibits phosphorylation by PNK and so forth). When you rinse with 70% EtOH you are simply diluting away those trace amounts of salt (while the precipitated sodium salt of DNA remains insoluble).
Thanks everybody, especially Damian and Paul sir for your more precise answer....it was very helpful
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