Experiments on purification of nucleic acids by ethanol precipitation with sodium acetate
Experiments on purification of nucleic acids by ethanol precipitation with sodium acetate
Nucleic acids are water-soluble compounds of polyanions that can combine with many 1-valent and 2-valent ions to form salts, the latter of which do not dissolve or become invariant in organic solvents. The latter is insoluble and invariant in organic solvents. It forms a precipitate at lower temperatures. This experiment is from the Mudanjiang Medical College undergraduate 5-year laboratory guide for testing majors.
Operation method
Experiments on purification of nucleic acids by ethanol precipitation with sodium acetate
Principle
Nucleic acids are water-soluble compounds of polyanions that can combine with many 1-valent and 2-valent ions to form salts, the latter of which do not dissolve or become invariant in organic solvents. They form precipitates at lower temperatures.
Materials and Instruments
Sample to be purified Move Experimental reagents: For more product details, please visit Aladdin Scientific website.
Sodium acetate buffer Anhydrous ethanol 70% ethanol
1. 3 mol / L sodium acetate buffer, pH 5.2: weighing sodium acetate (NaAC-3H20) 408.1 g, due to 800 ml of water, adjusted with glacial acetic acid to pH 5.2, plus distilled water to 1000 ml. divided and autoclaved (hereinafter referred to as 3 mol / L sodium acetate buffer).
2. Anhydrous ethanol and 70% (V/V) ethanol.
Experimental operation:
1. Transfer the solution containing nucleic acids to a new Eppendorf centrifuge tube with a pipette and measure the volume of the solution. 2.
2. Add 1/10 volume of 3 mol/L sodium acetate buffer. Make the final concentration of sodium acetate 0.3 mo1/L.
3. After inverted mixing, accurately add 2 times the volume (2.5 times the volume of RNA) of pre-cooled anhydrous ethanol, inverted mixing, and placed in an ice bath for 15-30 min or a 20 ℃ for 30 min.
4. 12000 r/min at 4℃, centrifuge for 15 min.
5. Hold the Eppendorf at an angle of 45 degrees so that the nucleic acid precipitate is facing upwards, and do not touch the precipitate with a micropipette or a pipette tip connected to a vacuum pump.
6. Add pre-cooled 70% ethanol to 2/3 of the volume of the tube. Mix and rinse to remove residual salts.
7. Centrifuge at 12000 r/min at 4℃ for 2 min and aspirate the supernatant as in step 5.
8. Leave the tube uncovered at room temperature for 10-l5min or in vacuum for 2 min to volatilize the ethanol.