TABLE 1.
Bacterial strains and plasmids
| Strain, plasmid, or primer | Characteristics or sequence (5′ to 3′) | Source or reference |
|---|---|---|
| Bacterial strains | ||
| M. bovis BCG | Pasteur strain; 1173P2 isolate | 13 |
| E. coli XL1 Blue | F′::Tn10proA+B+laclqΔ(lacZ)M15/recA1endA1gyrA96(NalR)thihsdR17(rk−mk+)supE44relA1lac | 7 |
| Plasmids | ||
| pTG627 | E. coli vector containing full-length SIVmac251 gag(p55) and pol genes | Transgene |
| pSN24 | E. coli/mycobacterial shuttle vector containing full-length SIVmac251 nef gene | 36 |
| pJN30 | E. coli/mycobacterial replicative shuttle vector; contains the lacZ gene from E. coli under control of up-regulated pBlaF* promoter from M. fortuitum and the transcription terminator of bacteriophage T4; contains kanR gene from Tn903 | 33 |
| pAV6950 | E. coli replicative/mycobacterial integrative shuttle vector; contains the attachment site and the gene coding for integrase of mycobacteriophage Ms6; contains kanR gene from Tn5 | 12 |
| pOIP23 | pAV6950 derivative; contains two transcription terminators of bacteriophage T4 around the unique EcoRI cloning site | This study |
| pJL17 | pJN30 derivative; contains entire nef gene of SIVmac251 under the control of pBlaF* (pBlaF*-nef cassette) | This study |
| pJL24 | pJN30 derivative; contains entire gag(p26) gene of SIVmac251 under the control of pBlaF* (pBlaF*-gag cassette) | This study |
| pJLN29 | pJN30 derivative; contains entire nef and gag(p26) genes of SIVmac251 organized as an operon under the control of pBlaF* (pBlaF*-nef-gag cassette) | This study |
| pOIP4 | pAV6950 derivative; contains pBlaF*-nef-gag cassette; sense orientation | This study |
| pOIP12 | pAV6950 derivative; contains pBlaF*-nef-gag cassette; antisense orientation | This study |
| pOIP33 | pOIP23 derivative; contains pBlaF*-nef-gag cassette isolated by two transcription terminators | This study |
| Primers | ||
| Kmr gene Tn5 | ||
| K1 | AGGTCTGCCTCGTGAAGAAGG | |
| K2 | CGTTGTGTCTCAAAATCTCTGA | |
| Kmr gene Tn903 | ||
| K3 | GGGCGAAGAACTCCAGCATG | |
| K4 | CAAGAGACAGGATGAGGATCG | |
| SIVmac251 nef gene | ||
| N1 | CGGTCCAAGCCGGCTGGA | |
| N2 | AATCCCTTCCAGTCCCCCC | |
| N3 | CCAAGTGGGATGACCCTTGGG | |
| JYL6 | TAGAATTCGGTACCTTAGCCAGAAGGAGAAGTACCGATGGGTGGAGCTATTTCCATG | |
| JYL7 | CCCAAGCTTGATATCGCTAGCTTAGCCTTCTTCTAACCTCTT | |
| SIVmac251 gag gene | ||
| G1 | CCGAGAACATTAAATGCCTGG | |
| G2 | TGATCCTGACGGCTCCCT | |
| G3 | GGCACTACTTCTGCTCCAAATTT | |
| JYL8 | TAGAATTCGCTAGCTAAGGTACCAGAAGGAGAAGTACCGATGGAAACTATGCCAAAAACAAG | |
| JYL10 | CCCAAGCTTGATATCTTATCTAGCCTTCTGTCCTGG |