Semi-dry electroblotting of DNA.
A procedure is described for the rapid transfer of DNA from agarose gels to nylon membranes using the semi-dry electroblotting technique. A Hind III digest of lambda DNA which was separated in a 1% agarose gel containing Tris, Borate, and EDTA (pH 8.0) was employed for the electrotransfer experiments. Transfer efficiency was determined by staining the DNA on the nylon membranes with a colloidal iron reagent. Current densities of 3-5 mA/sq. cm of gel permitted the transfer of high (23 kb) and low (0.3 kb) molecular weight fragments within 15 min. However, efficient transfer required a high ionic strength buffer that would prevent uneven dehydration of the agarose gel. Critical parameters for the transfer of nucleic acids with the semi-dry technique are discussed.
Fleming
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JE
Ryoichi Sasakawa Center for Aging Research, Linus Pauling Institute of Science and Medicine, Palo Alto, CA 94306.
Paull
T T
TT
D010582:Bacteriophage lambda / Q000032:analysis; D001709:Biotechnology; D002021:Buffers; D004279:DNA, Viral / Q000302:isolation & purification*; D004586:Electrophoresis / Q000379:methods*; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D004720:Endonucleases; D008567:Membranes, Artificial; D008970:Molecular Weight; D009757:Nylons
D002021:Buffers; D004279:DNA, Viral; D008567:Membranes, Artificial; D009757:Nylons; D004720:Endonucleases