Table 1 Primers used for cloning and site-directed mutagenesis
From: Identification of the feline foamy virus Bet domain essential for APOBEC3 counteraction
Name | Sequence (5′-3′ direction) |
|---|---|
Cloning of N-terminal deletion mutants of FFV Bet | |
FFVbel2ORF | ATCCCAAGCTTGCCACCATGGTCGGAAAGAATCCGGAAC (HindIII) |
FFVbel2ATG | ATCCCAAGCTTGCCACCATGGCTTGGGACAACCCTC (HindIII) |
FFVBet∆N82 | ATATCCCAAGCTTGCCACCATGGTGGTCACACGTCTGGTG (HindIII) |
FFVBet∆N92 | ATATCCCAAGCTTGCCACCATGGAATCATGGAAGAAGTATC (HindIII) |
FFVbel2as | GGGTCACTCGAGCTATTCAGAGTCAGATGACTC (SmaI) |
Cloning of V5 tagged FFV Bet and C-terminal deletion mutants of Bet | |
Bet-sense | CTCCCCTCGAAGCTTTCTGGGATATGTAAAACC (HindIII) |
dC1-V5 | GGGTCAGGTGCTGTCCAGGCCCAGCAGGGGGTTGGGGATGGGCTTGCCATCATCAGCTTGTGCTCTCC (SmaI) |
dC2-V5 | GGGTCAGGTGCTGTCCAGGCCCAGCAGGGGGTTGGGGATGGGCTTGCCCAGCAGAGAGTATTCTCCTC (SmaI) |
dC3-V5 | GGGTCAGGTGCTGTCCAGGCCCAGCAGGGGGTTGGGGATGGGCTTGCCTCTTCCATCAGGAAGTATCAC (SmaI) |
Bet-V5 antisense | GGGTCAGGTGCTGTCCAGGCCCAGCAGGGGGTTGGGGATGGGCTTGCCTTCAGAGTCAGATGACTCAG (SmaI) |
Alanine scanning mutgenesis | |
BetAlaRI | ATGCAAGATGATGGTAC C GCAGCGGCTCTAGTTAGCATAGTCAAATC (KpnI) |
BetAlaRII | ATGCAAGATGATGGTACCAACAATTCGGCAGCGGCCATAGTCAAATCCCTCTC (KpnI) |
BetAlaRIII | ATGCAAGATGATGGTACCACAATTCTCTAGTTAGGGCAGCGGCATCCCTCTCCCCACAATC (KpnI) |
BetAlaFI | ATGCAAGATGATGGTACCAGCCGCTGCCCTGGTACAAATCGCCGCTAC (KpnI) |
BetAlaFII | ATGCAAGATGATGGTACCAGGAATAGGCGCCGCTGCCATCGCCGCTACACTTAC (KpnI) |
BetAlaFIII | ATGCAAGATGATGGTACCAGGAATAGGCCTGGTACAAGCCGCCGCTACACTTACTAAAACC (KpnI) |
BetAlaR2 | AGGTAAAAGATTCCTATGTCTACGCACAATC (AccI) |
R-XhoI | ATGCAAGATGATGGTACCAACAATTCTCGAGTTAGCATAGTC (KpnI) |
ForwardVP | 5′ATGCAAGATGATCTCGAGAATTGTTGGCTGCCGGAATAGGCCTGGTAC (XhoI) |
ForwardV | 5′ATGCAAGATGATCTCGAGAATTGTTGGCCCCAGGAATAGGCCTGGTAC (XhoI) |
ForwardP | ATGCAAGATGATCTCGAGAATTGTTGGTAGCCGGAATAGGCCTGGTAC (XhoI) |
dGPL-AS1 | TGGAAGGGCAGCGGCGGGAACATCCTGCTTCTTG |
dGPL-S2 | GCCGCTGCCCTTCCAATTCTGAGTCCG |
dLPI-AS1 | ACTCAGGGCAGCGGCGAGTGGGCCGGGAACATCC |
dLPI-S2 | GCCGCTGCCCTGAGTCCGTATGTAATGG |
dLSP-AS1 | TACATAGGCAGCGGCAATTGGAAGGAGTGG |
dLSP-S2 | GCCGCTGCCTATGTAATGGCTTGGGACAACC |
dYVM-AS1 | CCAAGCGGCAGCGGCCGGACTCAGAATTGGAAG |
dYVM-S2 | GCCGCTGCCGCTTGGGACAACCCTCAG |
dDPD-AS1 | ATCACAGGCAGCGGCGGTTCTAGAACCTGTAATAC |
dDPD-S2 | GCCGCTGCCTGTGATCCTTTGTTCTGTAAG |
dCDP-AS1 | GAACAAGGCAGCGGCATCTGGGTCGGTTCTAGAAC |
dCDP-S2 | GCCGCTGCCTTGTTCTGTAAGTTGTTATGC |
dLFC-AS1 | CAACTTGGCAGCGGCAGGATCACAATCTGGGTCGGTTC |
dLFC-S2 | GCCGCTGCCAAGTTGTTATGCTGGAAAC |
dKLL-AS1 | CCAGCAGGCAGCGGCACAGAACAAAGGATCACAATC |
dKLL-S2 | GCCGCTGCCTGCTGGAAACAAAATATAC |
dNSE-AS1 | CCTCTCGGCAGCGGCCCCAGAGGCACTTCCAAATATG |
dNSE-S2 | GCCGCTGCCGAGAGGGTGTCATGGGCCAAAG |
dERV-AS1 | CCATGAGGCAGCGGCCTCACTGTTCCCAGAGGCAC |
dERV-S2 | GCCGCTGCCTCATGGGCCAAAGAGAATTC |
dSW-AS1 | CTCTTTGGCAGCGGCCACCCTCTCCTCACTGTTCC |
dSW-S2 | GCCGCTGCCAAAGAGAATTCTCACAGAG |
AS2-XmaI | AGTGTAAGTTCACCCGGGTCACTCGAGCTATTCAGAGTCAGATGACTC (SmaI, XmaI) |
Cloning of chimeric FFV/PFV Bet fusion proteins | |
chimera-s1 | ATGCAAGATGATAAGCTTTAGCTGCAGCAACAAAG (HindIII) |
ch1-as1 | GTGTTCCGGATTCTTTCCAGCAATAGTCAGCTCTCC |
ch1-s2 | GGAGAGCTGACTATTGCTGGAAAGAATCCGGAACAC |
ch2-as1 | GTGTTCCGGATTCTTTCCCTTCTGAGCAATCATTTC |
ch2-s2 | GAAATGATTGCTCAGAAGGGAAAGAATCCGGAACAC |
ch4-as1 | CCACGTGTGACCACGTTTGCATAGTGATCCTGGCTC |
ch4-s2 | GAGCCAGGATCACTATGCAAACGTGGTCACACGTGG |
ch5-as1 | GTAAGTGTAGCGGCGATCTGAATGTTCACCTGACC |
ch5-s2 | GGTCAGGTGAACATTCAGATCGCCGCTACACTTAC |
ch6-as1 | CAGGCACCATTCTTCTAGTTGCTTTTGGCCCATTGC |
ch6-s2 | GCAATGGGCCAAAAGCAACTAGAAGAATGGTGCCTG |
ch3-as1 | GTGTTCCGGATTCTTTCCCATTTCCTCTGGTGTGGGGATCC |
ch3-s2 | GGATCCCCACACCAGAGGAAATGGGAAAGAATCCGGAACAC |
ch-sense1- bspei | ATGCAAGATGATTCCGGAACACCCAAGACGGATC (BspEI) |
ch7-as1 | GATTCCAAAGAGGGTTGGCTGAGGGTTGTCCCAAGC |
ch7-s2 | GCTTGGGACAACCCTCAGCCAACCCTCTTTGGAATC |
ch8-as1 | CTGATAATTCTTATAAAATTGTACCAGGCCTATTCC |
ch8-s2 | GGAATAGGCCTGGTACAATTTTATAAGAATTATCAG |
ch9-as1 | GTCTGCACAGCCAGGTTTTTTGGTTACACTCTCTAGGGTC |
ch9-s2 | GACCCTAGAGAGTGTAACCAAAAAACCTGGCTGTGCAGAC |
ch10-as1 | AATCCTAGGATTGGTGAAGACTCTGGGATACAGGGAAGC |
ch10-s2 | GCTTCCCTGTATCCCAGAGTCTTCACCAATCCTAGGATT |
ch11-as1 | CTGTCAATGTTCTGATCTTCTCAGGATCACAGGCTATG |
ch1-s2 | CATAGCCTGTGATCCTGAGAAGATCAGAACATTGACAG |
chimera-as | GGGTCAGGTGCTGTCCAGGCCCAGCAGGGGGTTGGGGATGGGCTTGCCGAAGGGTCCATCTGAGTC (SmaI) |