Question

absorption of DNA to the purification column depends on various parameters. What are the two most important parameters to achieve maximum binding DNA to the purification column?

Answer 1

• As the binding of DNA to silica in the purification column is driven by dehydration and hydrogen bond formation, which competes against weak electrostatic repulsion, therefore, a high concentration of chaotropic salt helps to drive DNA adsorption onto silica. This ensures, proper adsorption of DNA as in case of a low concentration of salts, the DNA gets released.
• Also, the pH conditions act as an important parameter for determining the binding properties of DNA to the column. Furthermore, the removal of contaminants ( proteins, lipids, detergents and reagents used during cell lysis) is absolutely necessary to enable efficient binding to the purification column. Therefore, along with the binding buffer, chilled ethanol is added to precipitate the insolutble DNA and aggregate it , freeing it from all other immpurities.

Finally, if the pH and salt concentration of the binding solution are optimal, the nucleic acid binds to the silica gel membrane as the solution passes through the minicolumn purifcation column ( spin-based). Thus, for achieving maximum binding of DNA, the salt concentrationa nd pH conditions act as the most important parameters.