Infectious cDNA clone of North American porcine reproductive and respiratory syndrome (PRRS) virus and uses thereof
View Patent ↗The invention provides isolated polynucleotide molecules, including plasmids; viral vectors; and transfected host cells that comprise a DNA sequence encoding an infectious RNA sequence encoding a North American PRRS virus; and also North American PRRS viruses encoded thereby. The invention further provides isolated infectious RNA molecules encoding a North American PRRS virus. The invention also provides isolated polynucleotide molecules, infectious RNA molecules, viral vectors, and transfected host cells encoding genetically-modified North American PRRS viruses; and genetically-modified North American PRRS viruses encoded thereby. The invention also provides vaccines comprising such plasmids, RNA molecules, viral vectors, and North American PRRS viruses, and methods of using these vaccines in swine and in other animals. Also provided are isolated polynucleotide molecules, viral vectors, and transfected host cells that comprise a nucleotide sequence encoding a peptide of a North American PRRS virus. These viral vectors and transfected host cell lines are useful in providing peptides to compensate for mutated peptide coding sequences of DNA sequences encoding genetically-modified North American PRRS viruses so that functional virions can be generated.
1. An isolated polynucleotide molecule comprising a DNA sequence encoding an infectious RNA molecule encoding a North American PRRS virus, wherein said DNA sequence is SEQ ID NO:1, or a sequence that has at least 70% identity to SEQ ID NO:1, or that hybridizes to the complement of SEQ ID NO:1 under highly stringent conditions which comprise hybridization to filter bound DNA in 0.5M NaHPO4, 7% SDS, 1mM EDTA at 65° C., and washing in 0.1×SSC/0.1% SDS at 68° C., and wherein said isolated polynucleotide molecule further comprises one or more further nucleotide sequences inserted into the nonstructural protein 2 coding region of ORF1 a, each of which encodes a detectable heterologous antigenic epitope, wherein PRRS virus sequence is not deleted as a result of said insertion.
2. The isolated polynucleotide molecule of claim 1 , wherein said one or more nucleotide sequences that each encode a heterologous antigenic epitope are inserted in the hypervariable region (genomic nucleotides 2720-3980) in the nonstructural protein 2 coding region of ORF1a.
3. The isolated polynucleotide molecule of claim 2 , wherein a nucleotide sequences that encodes a heterologous antigenic epitope is inserted at position 3219 in the hypervariable region in the nonstructural protein 2 coding region of ORF1a.
4. The isolated polynucleotide molecules of claim 1 that is genetically modified such that when it infects a porcine animal it is unable to produce PRRS in the animal.
5. An isolated polynucleotide molecule according to claim 1 that is at least 75% identical to SEQ ID NO:1.
6. An isolated polynucleotide molecule according to claim 1 that is at least 85% identical to SEQ ID NO:1.