Table 1.

CODEHOP and gene-specific primers for cloning and expression of RV1 and RV2 rhadinovirus ORF59 homologs

Virus/[Sequence source]1	Host Species/[DNA source]	Primers (gene)2	Primer Sequence (5'-3')
RV1 Lineage
KSHV [U93872]	Human (H. sapiens) [BCBL cells]	KSHV ORF59a3 KSHV ORF59b3	GTACAAGGATCCCCTGTGGATTTTCACTATGG ACAGATAAGCTTAAATCAGGGGGTTAAATGTGG
PtrRV1 (also known as PanRHV1/PtRV1) [this study]	Chimpanzee (P. troglodyte) [Ptr001]	SRDEa (ORF60)4 PQFVb (ORF59)4 IGNGa (ORF59)5 PtrRV1 ORF59a3 PtrRV1 ORF59b3	CTGGCTAACGACTACATCTCCAGRGAYGARCT CCGTAAGAAATGGTGGTCCTGACRAAYTGNGG GAATACTTCCATCGGTAACG TATATAAGATCTAAATCAGTGGGTTAAATGTGG ATTATAAGATCTCCTGTGGATTTTCACTATGG
RFHVMn [this study]	Pig-tailed macaque (M. nemestrina) [Mne442N]	NFFEa (ORF60)5 PQFVb (ORF59)4 YGVRb (ORF59)5 WCFIb (ORF58)4 RFHVMn ORF59a3 RFHVMn ORF59b3	GGCAGTTTCAAGGCTGTGAATTTTTTTGAGCG CCGTAAGAAATGGTGGTCCTGACRAAYTGNGG CGTCCACCCTGACCCCATA CGAGTACAGGGCCTTGAAGATRAARCACCA GTACAAGGATCCCCTGTGGATTTTCATTATGG GAACTGAAGCTTTTAAATTAATGGGTTAAACG
RFHVMm [this study]	Rhesus macaque (M. mulatta) [MmuYN91]	NFFEa (ORF60)5 PQFVb (ORF59)4 FTHTa (ORF59)5 YYELb (ORF58)4 RFHVMn ORF59a3 RFHVMm ORF59b3	GGCAGTTTCAAGGCTGTGAATTTTTTTGAGCG CCGTAAGAAATGGTGGTCCTGACRAAYTGNGG CAACGGATTCACGCACACG AAATGCTCCGCAGAAGCCCAGYTCRTARTA GTACAAGGATCCCCTGTGGATTTTCATTATGG GAACTGAAGCTTTCAAATCAACGGGTTAAAGG
RFHVMf [this study]	Cynomolgus macaque (M. fascicularis) [Mfa95044]	EVEGa (ORF59)4 HRYYb (ORF58)4 NAAKb (ORF59)5 MPVDa (ORF60)5 YYELb (ORF58)4	TGGCACTCCAACGAAATATTAGARGTNGARGG TGCTAAAAATCCAAGTTCGTARTAYCTRTG CTTCGCAGCATTCCAGGAC ATCATGCCTGTGGATTTTCA AAATGCTCCGCAGAAGCCCAGYTCRTARTA
RV2 Lineage
PtrRV2 (also known as PanRHV2) [this study]	Chimpanzee (P. troglodyte) [Ptr001]	LYNTa (ORF60)4 EMFGb (ORF59)5 TREMa (ORF59)5 GTYTb ORF58)4 PtrRV2 ORF59a3 PtrRV2 ORF59b3	GGCCGCCGGCATGCTGTACAAYACNATGAT TACCCGTGAGATGTTTGGAG CCCATACCAGAGAAATGTTC GAGGGACGCCTCCGACGTGTNCGTNCCCAT ATCATAGATCTCCTATCACATTTCACTACGGAG ATATATAAGCTTAAATAAGGGGATTAAATGTAG
MneRV2 (also known as PRV/MGVMn) [this study]	Pig-tailed macaque (M. nemestrina) [Mne442N]	RDELa (ORF60)4 PQFVb (ORF59)4 EMFGa (ORF59)5 CFICb (ORF58)4 MneRV2 ORF59a3 MneRV2 ORF59b3 T1b (ORF59)6 T2b (ORF59)6 T3b (ORF59)6 T4b (ORF59)6 T5a (ORF59)7 T6a (ORF59)7	CTTGCCAACGATTACATTTCCAGRGAYGARCT CCGTAAGAAATGGTGGTCCTGACRAAYTGNGG TACCCGTGAGATGTTTGGAG TACAAAATACAGCGAGTGATANATRAARCA GTACAAGGATCCCCGGTCTCGTTCCACTACG TAACTGAAGCTTCTAAAACAGCGGGTTGAAGG CTAATTAAGCTTCTAAACTCCAAACATCTCACGGG CTAATTAAGCTTCTAGGTAAACGTGGCAACGGC CTAATTAAGCTTCTAGCCCAACTTGACGTCAGC CTAATTAAGCTTCTACGTTCGCGGTGATTTGGC GTACAAGGATCCCCTACCGGCCAGGAGAATG GTACAAGGATCCAAATCACCGCGAACGAACG
RRV 17577 [NC_003401]	Rhesus macaque (M. mulatta)	RRV ORF59a3 RRV ORF59b3	GTACAAGGATCCCCTGTCTCGTTTCATTACGG GAACTGAAGCTTAAAACAACGGGTTGAACG
MfaRV2 (also known as MGVMf) [this study]	Cynomolgus macaque (M. fascicularis) [Mfa98044]	RDELa (ORF60)4 NRb (ORF59)5 NRa (ORF59)5 CFICb (ORF58)4 MfaRV2 ORF59a3 MfaRV2 ORF59b3	CTTGCCAACGATTACATTTCCAGRGAYGARCT GGCCCGGAAAATGAGTAACA TCTGAATATGTCACATCCGTTCATA TACAAAATACAGCGAGTGATANATRAARCA GTACAAGGATCCCCTGTCTCGTTTCATTACGG TAACTGAAGCTTCTAAAACAGCGGGTTGAACG

¹ We have utilized the virus nomenclature scheme indicating the host genus (first letter), species (second and third letters) with the designation of rhadinovirus lineage, ie RV1 or RV2, to provide a unique designation for the rhadinoviruses from different Old World primate species. For historical reasons, Homo sapiens HsaRV1 remains as KSHV, the macaque RV1s remain as RFHVMn, RFHVMm, and RFHVMf and the rhesus macaque MmuRV2 remains as RRV. The gene-specific primers are derived from published sequences (Genbank accession number is indicated) or from new sequences obtained in this study.

² a and b designations indicate forward and reverse strand primers, respectively

³ 5' and 3' gene-specific primers used for preparing the full-length ORF59 bacterial expression vectors

⁴ CODEHOP PCR primers used for cloning the initial regions of the ORF59 genes

⁵ gene-specific primers used in conjunction with the CODEHOP PCR primers to obtain the full-length ORF59 coding sequences

⁶ gene-specific primers used in conjunction with the MneRV2 ORF59a primer to construct MneRV2 ORF59 truncation mutants

⁷ gene-specific primers used in conjunction with the MneRV2 ORF59b primer to construct MneRV2 ORF59 truncation mutants