Construction of a cloned library of adenovirus DNA fragments in bacteriophage M13.
The construction of recombinant M13 phages containing adenovirus DNA inserts was undertaken to provide strand-specific hybridization probes for analyses of adenovirus type 2 RNA transcripts. A library of molecular probes was constructed by cloning restriction endonuclease fragments of adenovirus types 2 and 5 DNA in the duplex replicative form DNA of the single-stranded bacteriophage vectors, M13mp7, M13mp8, and M13mp9 (Messing, J., and Vieira, J. (1982) Gene 19,269-276). Adenovirus DNA segments from early, intermediate, and late gene regions, accounting for at least 95% of the adenovirus chromosome, have been cloned in both possible orientations using these M13 derivatives as vectors. DNA cloned into these vectors can readily be obtained in a circular single-stranded form directly from mature phage particles. The cloned DNA fragments have been oriented and further characterized by restriction endonuclease mapping and hybridization with 32P-labeled adenovirus DNA. The polarity and fidelity of the adenovirus DNA in the recombinant phages has been confirmed by hybridization with labeled adenovirus 2 early and late mRNA. Restriction endonuclease analyses of M13 clones containing adenovirus DNA inserts spanning genome coordinates 31.7-56.9 have indicated that the relative locations of some restriction coordinates located within this region do not correspond to the mapped restriction sites in the DNA of adenovirus 2. Potential uses for these M13 clones in studies of adenovirus gene expression are discussed.
D000256:Adenoviridae / Q000235:genetics*; D001435:Bacteriophages / Q000235:genetics*; D004262:DNA Restriction Enzymes / Q000378:metabolism; D004274:DNA, Recombinant / Q000032:analysis; D004279:DNA, Viral / Q000032:analysis*; D009693:Nucleic Acid Hybridization; D014158:Transcription, Genetic
D004274:DNA, Recombinant; D004279:DNA, Viral; D004262:DNA Restriction Enzymes