Alkaline Blotting: The Procedure

Positively charged nylon membranes allow the use of alkaline transfer buffers , which link the nucleic acids to the membrane without UV crosslinking. In some cases, alkaline blotting gives a higher background. Increasing the concentration of blocking reagent will often elimintate this problem.

NORTHERN BLOTTING

Follow the basic procedure given in the standard protocol for Northern Blotting, using 40mM NaOH in place of the 10X SSC.

SOUTHERN BLOTTING

Follow the procedure in the standard protocol for Southern Blotting, omitting Step 5 and using 0.4M NaOH in place of the 10X SSC.