Table 5.
Description of strains, plasmids, and primers used in this study.
| Plasmid, Primer, or Strain | Description or Sequence | Source or Reference |
|---|---|---|
| F. tularensis strains | ||
| LVS | F. tularensis subsp. holarctica live vaccine strain | Karen Elkins |
| RZR46 | Spontaneous resazurin-resistant LVS mutant containing deletion of single thymine residue at position 2 in dipA | This study |
| RZR47 | Spontaneous resazurin-resistant LVS mutant containing 532G > T substitution in pilD | This study |
| ΔdipA | LVS dipA deletion mutant | This study |
| FTL_0959d | LVS pilD disruption mutant | This study |
| E. coli strains | ||
| DH5α | fhuA2 Δ(argF-lacZ)U169 phoA glnV44 Φ80 Δ(lacZ)M15 gyrA96 recA1 relA1 endA1 thi-1 hsdR17 | NEB 1 |
| Plasmids | ||
| pABST | pFNLTP8 with F. tularensis LVS groE promoter | [11] |
| pDipA | pABST containing dipA under control of groE promoter | This study |
| pPilD | pABST containing dipA under control of groE promoter | This study |
| pJH1 | pMQ225 with the I-SceI restriction site | [26] |
| pGUTS | pGRP with I-SceI under the control of FGRp | [26] |
| pJH1ΔdipA | pJH1 with the left and right 500-bp flanking regions of dipA cloned into the PstI-SphI site of pJH1 | This study |
| pJH1-0959d | pJH1 with the central 470 base pair region of FTL_0959 | This study |
| Primers | ||
| 1F_dipA | 5′-CATGCTGCAGGTAGTGTTTGGATCTTTTGTATTAGCAG -3′ | IDT 2 This study |
| 2R_dipA | 5′-ATAATAAGTCGACGGTACCACCGGTAATTATAAATTTAGCTTTCATCTATTTCTCCTG-3′ | IDT This study |
| 3F_dipA | 5′-ACCGGTGGTACCGTCGACTTATTATTACTACCAAAGAGCAGCAAAG-3′ | IDT This study |
| 4R_dipA | 5′-CATGGCATGCCATCACCACATTTAGAACTATTGGC-3′ | IDT This study |
| F_0959d | 5′-CATGGGATCCGGGATTGATAAGCAACAATCTCCAC-3′ | IDT This study |
| R_0959d | 5′-CATTGCATGCCCAAAGCCTTCTTTACCTGTAAGG-3′ | IDT This study |
| dipA_F | 5′-CATGGAATTC GTGACGCTTGATGTTTTTGTATTGCAGG-3′ | IDT This study |
| dipA_R | 5′-CATGGCTAGCGCGCTATTTAGAAGTCACCGCATTTTG-3′ | IDT This study |
| pilD_F | 5′-ATCGGAATTCATGTTGTTATATATCGAGTGCCAAATAAAC-3′ | IDT This study |
| pilD_R | 5′-ATCACATATGTTATACATAGATATTATCTTTAGTCAGTAG ATAAAAAAATGTTG-3′ | IDT This study |
| pJH1_conf | 5′-CTGATTTAATCTGTATCAGGCTGAAAATC-3′ | IDT This study |
1 NEB—New England Biotechnologies. 2 IDT—Integrated DNA Technologies.