Preparation of Genomic Blots

1. Digest high-molecular-weight genomic DNA (10-15 |g) with various restriction enzymes (100 U) (see Note 3) in 200 |L of appropriate buffers. Aliquots of the digest should be run through agarose gels to make sure that the digestion is complete, as well as to estimate the concentration of genomic DNA in each solution.

2. Ethanol precipitate the digested genomic DNA and redissolve in 30 |L of TE. Load the solution onto agarose gels and run through 0.8% agarose gels overnight. For better resolution, electrophoresis at 2 V/cm gel length is recommended. It is important that each lane contains equal amounts of genomic DNA for the comparison of signal intensities between lanes.

3. Transfer to a nitrocellulose membrane by a standard procedure of Southern blotting.

4. Bake the nitrocellulose membrane at 80°C in a vacuum over for 2 h.

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