TABLE 1.
Strains and plasmids used in this study
| Strain | Subspecies/alternative strain designation(s)/genotype/phenotype | Source/reference |
|---|---|---|
| F. tularensisa | ||
| FSC155 (LVS1) | F. tularensis subsp. holarctica, live vaccine strain, LVS (in strain collection since 1993) | ATCC 29684, |
| FSC458 (LVS2) | Past tularemia vaccine, code no. NDBR 101, lot no. 11; vial reconstituted and used, in strain collection since June 2004 | The National Drug Company |
| FSC200 | F. tularensis subsp. holarctica | Human ulcer, 1998, Sweden |
| FSC352 | F. tularensis subsp. holarctica | Hare, 1974, Sweden |
| FSC237 | F. tularensis subsp. tularensis, SCHU S4 | Human ulcer, 1941, Ohio |
| FSC472 | FSC155/pAL10, FSC155 expressing pilA in trans, Tcr | This study |
| FSC681 | FSC458/pAL10, FSC458 expressing pilA in trans, Tcr | This study |
| FSC688 | FSC458/pilA+, FSC458 expressing pilA in cis | This study |
| FSC741 | FSC458/FTT0086+, FSC458 expressing FTT0086 in cis | This study |
| FSC693 | FSC458/FTT0918+, FSC458 expressing FTT0918 in cis | This study |
| FSC694 | FSC458/pilA+/FTT0918+, FSC458 expressing pilA and FTT0918 in cis | This study |
| FSC767 | FSC200 /FTT0918−, FTT0918 insertion mutant | This study |
| E. coli | ||
| Top10 | F−mcrA Δ(mrr-hsdRMS-mcrBC), φ80lacZΔM15 ΔlacX74 recA1 deoR araD139 (Δara-leu)7697 galU galK rpsL (Smr) endA1 nupG | Invitrogen |
| S17-1λpir | recA thi pro hsdR−M+, <RP4:2-Tc:Mu:Km:Tn7>Tpr Smr | 32 |
| BL21 Star (DE3) | F−ompT hsdSB (rB− mB−) gal dcm rne-131 (DE3) | Invitrogen |
| Plasmids | ||
| pCR2.1-TOPO | Topo cloning vector, Ampr Kmr | Invitrogen |
| pCR4.0-TOPO | Topo cloning vector, Ampr Kmr | Invitrogen |
| pKK214GFP | groELS promoter, Ft ori, p15A ori, GFP,b Tcr | 1 |
| pDM4 | Suicide plasmid. sacB, mobRP4, oriR6K, Cmr | 21 |
| pDMK2 | Suicide plasmid. sacB, mobRP4, oriR6K, Kmr | K. Kadzhaev, unpublished |
| pFTKan1 | EZ-Tn5, pMOD-2 F. tularensis codon-optimized KanR clone sequence, F. tularensis groELS promoter | T. Kawula, unpublished |
| pET-30a(+) | Cloning vector carrying an N-terminal His-tag/thrombin/S-tag/enterokinase configuration and an optional C-terminal His tag sequence, Kmr | Novagen |
| pAL10 | pilA gene cloned into NdeI and EcoRI sites of pKK214GFP, Tcr | 7 |
| pAL11 | pilA gene and additionally 1 kb upstream and downstream of pilA cloned into XhoI and SacI sites of pDM4, Cmr | 7 |
| pEMS36 | FTT0086 gene and additionally 1 kb upstream and downstream of FTT0086 cloned into the XhoI and SalI sites of pDM4, Cmr | This study |
| pKK223 | The cis complementation construct for FTT0918 cloned into SaII and NotI sites of pDMK2, Kmr | This study |
| pKK224 | PvuII fragment generated by PCR using primers groELS/KanR/E-C cloned into blunt-ended pDM4, Cmr | This study |
a
All Francisella strains are from the Francisella Strain Collection at FOI, Umeå, and each strain has been given a strain collection number.
b
GFP, green fluorescent protein.