Fig. Different steps of genomic DNA prepration.

General Steps for preparation of genomic DNA:

1.   Disruption

2.   Lysis

3.   Removal of protein and other contaminants

4.  Recovery of DNA

Note: In some methods step 1 and 2 are combined.

Crude lysate preparation:

DNA can be purified very simply by incubating cell lysate at higher temperature (95°C, 20 min.) or to treate the cell lysate with proteinase K and the product can be directly used for further applications. This method is comparatively very simple and straight forward but, the quality of purified DNA will be poor and can be useful for few applications only. The DNA isolated with this method will not be at optimum pH and get degraded during storage, further more incomplete inactivation of proteinase K results several complications. The DNA will be contaminated with salts and several other compounds which can inhibit the enzyme activity in downstream process.

DNA extraction with salts:

The salting out phenomenon can be used for DNA isolation. The crude extract can be directly treated with high salt (potassium acetate or ammonium acetate) concentration for precipitation of proteins and other contaminants. The precipitates can be removed by centrifugation, and the supernatant can be further treated with alcohol for DNA precipitation. Removal of proteins and other contaminants using this method may be inefficient, and RNase treatment, dialysis, and/or repeated alcohol precipitation are often necessary before the DNA can be used in downstream applications. DNA yield and purity are highly variable using this method.