Lower temperatures promote the flocculation of the nucleic acids so they form larger complexes that pellet under the centrifugal forces of a microcentrifuge. A nucleic acid concentration high enough to force the DNA out of solution if the concentration is not high enough, you can add a carrier nucleic acid or glycogen to enhance the recovery. DNA is less soluble in isopropanol so it precipitates faster even at low concentrations.
With ethanol, the DNA needs to be at a higher concentration to flocculate but the salt tends to stay soluble, even at colder temperatures.
So for the typical precipitation protocol, isopropanol is added from between 0. If you are precipitating small volumes of DNA, and you can fit the required amount of solvent into the sample tube, then ice-cold ethanol is the preferred choice.
Isopropanol is useful for large sample volumes e. Because less isopropanol is needed for precipitation, you can often fit your sample and the solvent in one 15 ml tube. However, because salts are generally less soluble in isopropanol than in ethanol, they tend to co-precipitate with DNA. To minimize the likelihood of salt precipitation, isopropanol precipitation is best at room temperature with short incubation times. Because DNA is less soluble in isopropanol, isopropanol allows precipitation of larger species and lower concentrations of nucleic acids than ethanol, especially if you incubate at low temperatures for long periods of time.
So now you know the difference between ethanol and isopropanol precipitation, and when to use each method. Good luck with your DNA precipitations!
Has this helped you? Then please share with your network. Hi in my script it is nowhere mentioned that we are supposed to use salt. Does it still work without adding salt or is it a mistake in the script? In addition, isopropanol is often used for precipitating DNA from large volumes as less alcohol is used see protocols below.
The ethanol and isopropanol can also wash away the remaining salt residue. After being washed in alcohol and subjected to a centrifuge, the precipitated DNA protein will form a pellet, which can be washed in alcohol again, dried, and re-suspended in a Tris or TE buffer.
Be careful not to overdry the sample, since this can denature the DNA; just leave the washed pellet on the lab table for a few minutes. If isopropanol has been used during the extraction instead of ethanol, the sample may not adhere as tightly to the tube and may require a longer drying time. Incubate on ice for 15 minutes. In case of small DNA fragments or high dilutions overnight incubation gives best results. Discard supernatant by decanting or pipetting, being careful not to throw out DNA pellet which may or may not be visible.
Add 0. Calculate after addition of sodium acetate. Room temperature isopropanol minimizes coprecipitation of salt. Discard supernatant by decanting, being careful not to throw out DNA pellet which may or may not be visible. Isopropanol precipitated pellets are often difficult to see and loosely attached.
Mark outside of tube before centrifugation for easy identification. Topics: Molecular Biology. This assay is suitable for the simple and rapid estimation of protein concentration. This assay is based on a single Coomassie dye based reagent.
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