Key Points
- To clone the RNA genome of poliovirus type 1 into bacterial plasmids and construct a comprehensive physical restriction map.
- Synthesized cDNA-RNA hybrids from the poliovirus type 1 Mahoney strain using reverse transcriptase and inserted them into Escherichia coli plasmid pBR322.
- Identified recombinant colonies through in situ hybridization with virus-specific RNase T1-resistant oligonucleotides.
- Characterized cloned cDNA inserts using restriction endonuclease mapping and electron microscopy.
- Isolated overlapping cDNA clones up to 3.2 kilobase pairs in length, spanning at least 99% of the viral RNA genome.
- Reconstructed the complete restriction map of the poliovirus genome from overlapping fragments.
- Cloned the complete 5' end of the viral genome within recombinant plasmid pPV1-366.
Structured PICO
PPopulationMahoney strain of poliovirus type 1
IInterventionMolecular cloning using reverse transcriptase and Escherichia coli plasmid pBR322
OOutcomeSuccessful cloning and restriction mapping of the poliovirus genome
The study successfully cloned nearly the entire genome of poliovirus type 1, enabling detailed restriction mapping and further molecular analysis.