A METHOD FOR the DETERMINATION of STRANDEDNESS of DNA FRAGMENTS CLONED IN PHAGE M13 |
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| Authors: | SAUMYA BHADURI |
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| Affiliation: | U.S. Department of Agriculture, ARS Eastern Regional Research Center 600 E. Mermaid Lane Philadelphia, PA 19118 |
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| Abstract: | Recombinant M13Hol phage containing Eco R1 restriction endonuclease fragments B, E, and F of adenovirus type 2 (Ad2) DNA were constructed by cloning into the unique Eco R1 site of the replicative form of the phage M13Hol176 DNA. Polarity of the adenovirus inserts in recombinant molecules was deduced by the following procedures: Viral DNA fragments obtained from Ad2 DNA molecules were purified, denatured, and subjected to electrophoresis. the separated DNA strands were transferred from agarose to nitrocellulose by the Southern procedure and hybridized with radioactive 3'-end labeled Hae III fragments of the recombinant phage DNAs. This procedure provided a rapid test for assaying strandedness of the cloned fragments. |
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