EFFICIENT RECOVERY OF CLONED HUMAN CYTOMEGALOVIRUS DNA FRAGMENTS FROMAGAROSE GELS

A simple and efficient method for the recovery of DNA fragments from a garose gels is described. After electrophoresis, bands of interest are cut out of the gel and agarose slices pushed through the opening of a syringe needle. The resulting gel slurry is frozen and thawed three t imes and then centrifuged. DNA in the supernatant is precipitated, res uspended in a small volume of buffer and, finally, desalted. A recombi nant pAT153 plasmid carrying the BamHI-P fragment of the human cytomeg alovirus (HCMV) genome was detected using purified viral HindIII-E DNA fragment as a probe. Restriction endonuclease analysis was used to co nfirm the identity of the cloned fragment. Experiments performed with the recombinant pLCR127 plasmid indicate that our freeze/ thaw method, with about 80% recovery and very little DNA degradation, is more adva ntageous than the well known electroelution and NaI/glass methods.