Northern Blotting

Northern Blotting

Adapted from Molecular Cloning (Maniatis), Current Protocols in Molecular Biology, and J. Yost Lab (UMN)

Objective:

Northern blotting allows detection of specific RNA sequences. RNA is fractionated by agarose gel electorphoresis, followed by transfer (blotting) to a membrane support, followed by hybridization with DNA or RNA probes.

Procedures:

Start=> This protocol requires isolated total RNA or poly(A)+ RNA.

  1. You must first run the RNA on a denaturing gel; a formaldehyde or glyoxal/DMSO gel.
  2. Next, you need to transfer the RNA from the gel to a solid support membrane, preferably nylon.
  3. Once, the membrane with the immobilized RNA is available, hybridization with the appropriate probe is required.