Horizontal two-dimensional electrophoresis of complex DNA samples using precast gel systems.

We have developed a simplified procedure for the separation of enzyme-digested genomic DNA, or of complex mixtures of cloned DNA, into two dimensions. The procedure relies on the use of precast gels for horizontal electrophoretic separations. Precast agarose-type gels are used for the first-dimensional separation of fragments based on size. Precast polyacrylamide gels are used for the second-dimensional separation of fragments. The separated fragments are subjected to enzymatic digestion in situ prior to their transfer to the second-dimensional gel. Applications of this procedure include the analysis of DNA libraries, analysis of yeast artificial chromosomes (YACs) as well as other similar preparations, and the screening of genomic DNA for the occurrence of multi-copy DNA fragments as in the case of genomic amplifications in cancer.

Reduced chemical and radioactive liquid waste during electrophoresis using polymerized electrode gels.

Using polyacrylamide electrode gels during horizontal sodium lauryl sulfate-polyacrylamide electrophoresis of radiolabeled proteins instead of electrode papers reaching into electrode buffer reservoirs, silver staining is improved and we reduce chemical and radioactive liquid waste during electrophoresis.