Genomic cloning and restriction site mapping of a porcine adenovirus isolate: demonstration of genomic stability in porcine adenovirus.
Restriction endonuclease maps were constructed for the genome of a porcine adenovirus (PAV), NADC-1, which was isolated in 1972 from an adult swine. Genomic DNA libraries of NADC-1 Bam HI, Eco RI/Bam HI, and Sph I fragments were cloned into pUC-18. Using the cloned NADC-1 Bam HI and Eco RI/Bam HI fragments as probes, Southern blot hybridizations were performed to human adenovirus 2 (Ad-2) restriction fragments to determine the left-to-right orientation of the Bam HI and Eco RI/Bam HI fragments. Genomic NADC-1 DNA was cleaved with ten restriction endonucleases (RE). Using cloned NADC-1 genomic fragments as probes in Southern blot hybridizations, an RE site map was constructed. Nucleotide sequencing of four clones confirmed several RE sites. The size of the NADC-1 genome was determined to be approximately 32 kbp. The size of Hind III, Xba I, Sma I, Eco RI, Bam HI, Bgl II, Pst I, and Sph I RE fragments from NADC-1 was compared to those from the reference strain of PAV serotype 4 (F618), and to two recent isolates, NADC-2 and NADC-3. For all restriction enzymes examined, the sizes of the NADC-1 fragments were identical to PAV-4, NADC-2, and NADC-3 fragments, indicating that the NADC-1 isolate is very closely related, if not identical, to PAV-4 and two recent isolates. Southern blot hybridizations also indicated that NADC-1, NADC-2, NADC-3, and PAV-4 are very similar and revealed regions of sequence similarity between NADC-1 and human Ad-2 and human Ad-5.
Kleiboeker
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Virology Swine Research Unit, National Animal Disease Center, USDA, Ames, Iowa.
Seal
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Mengeling
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D000256:Adenoviridae / Q000235:genetics*; D000260:Adenoviruses, Human / Q000235:genetics; D000818:Animals; D015139:Blotting, Southern; D002460:Cell Line; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000032:analysis / Q000302:isolation & purification; D004587:Electrophoresis, Agar Gel; D012150:Polymorphism, Restriction Fragment Length; D015183:Restriction Mapping; D013552:Swine
D004279:DNA, Viral