One-hour downward alkaline capillary transfer for blotting of DNA and RNA.
The downward alkaline capillary transfer of DNA and RNA from agarose gel to a hybridization membrane was performed using a transfer solution containing 3 M NaCl and 8 mM NaOH. Under mild alkaline conditions, DNA and RNA were completely eluted from the agarose gel and bound to a hybridization membrane within 1 h. On the basis of this new method of transfer a blotting protocol, downward alkaline blotting, was elaborated. It provides a fast and efficient alternative to commonly used Southern and Northern blotting protocols. The downward alkaline blotting of DNA and RNA can be completed in 2.5 and 1.5 h, respectively, and can be used with both plastic and nitrocellulose membranes. In addition, the downward alkaline blotting protocol allows for a hybridization efficiency of DNA and RNA higher than that of the standard blotting protocols performed at neutral pH.
Chomczynski
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Division of Endocrinology, College of Medicine, University of Cincinnati, Ohio 45267.
D000818:Animals; D015152:Blotting, Northern / Q000379:methods*; D015139:Blotting, Southern / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D008567:Membranes, Artificial; D009693:Nucleic Acid Hybridization; D051381:Rats; D012965:Sodium Chloride; D012972:Sodium Hydroxide; D013997:Time Factors
D008567:Membranes, Artificial; D012965:Sodium Chloride; D012972:Sodium Hydroxide