How can I precipitate genomic DNA using isopropanol?
FAQ ID -2953

Alcohol precipitation is commonly used for concentrating, desalting, and recovering nucleic acids. Since less alcohol is required for isopropanol precipitation, this is the preferred method for precipitation of DNA from large volumes. In addition, isopropanol precipitation can be performed at room temperature, which minimizes co-precipitation of salt that interferes with downstream applications.

 

Procedure

  1. Adjust the salt concentration, for example, with sodium acetate (0.3 M, pH 5.2, final concentration) or ammonium acetate (2.0–2.5 M, final concentration).
  2. Add 0.6–0.7 volumes of room-temperature isopropanol to the DNA solution and mix well.
  3. Centrifuge the sample immediately at 10,000–15,000 x g for 15–30 min at 4°C
  4. Carefully decant the supernatant without disturbing the pellet.
  5. Wash the DNA pellet by adding 1–10 ml (depending on the size of the preparation) of room-temperature 70% ethanol. This removes co-precipitated salt and replaces the isopropanol with the more volatile ethanol, making the DNA easier to redissolve.
  6. Centrifuge at 10,000–15,000 x g for 5–15 min at 4°C.
  7. Carefully decant the supernatant without disturbing the pellet.
  8. Air-dry the pellet for 5–20 min (depending on the size of the pellet).
  9. Redissolve the DNA in a suitable buffer.

Tip: Use a buffer with a pH of 7.5–8.0, as DNA does not dissolve easily in acidic buffers. Often distilled water can have an acidic pH. The addition of EDTA protects the DNA from DNase digestion.

Tip: High-molecular-weight DNA, such as genomic DNA, should be redissolved very gently to avoid shearing. If the DNA pellet does not dissolve easily, heat at 55°C for 1–2 h with gentle shaking.