TABLE 1.
Plasmids and strains used in this study
| Plasmid or strain | Description | Antibiotic resistancea | Source or reference |
|---|---|---|---|
| Plasmids | |||
| pKSV7 | Temp-sensitive shuttle vector | Cb, Cm | 18 |
| pPL2 | Site-specific integration vector | Cm | 43 |
| pIMC3ery, pIMC3tet, pIMC3kan | Site-specific integration vectors with IPTG-induced expression of Ery, Tet, Kan | Cm, Ery, Tet, Kan | 44 |
| pAD1-cGFP | Phyper-driven expression of GFP (constitutive) in pPL2 derivative | Cm | 45 |
| pTM2 | InlAm subcloned into PstI/BamHI-digested pKSV7 | Cb, Cm | This study |
| pGJ-cGFP | Phelp from pIMC3 subcloned into SacI/EagI-digested pAD1-cGFP | Cm | This study |
| pAF1a | 0.98-kb fragment upstream of lplA1 (−988 to −1) in pKSV7 | Cb, Cm | This study |
| pAF1-1 | 1.042-kb fragment from bp 954 of lplA1 to bp +1001 downstream adjacent to 0.98-kb fragment upstream of lplA1 in pAF1a | Cb, Cm | This study |
| pTML1 | lplA1 plus 988 bp of upstream DNA from L. monocytogenes InlAm in SalI/PstI-digested pIMC3ery | Cm, Ery | This study |
| Bacterial strains | |||
| E. coli | |||
| DH5α | F− endA1 hsdR17(rK− mK−) supE44 thi-1 λ− recA1 gyrA96 relA1 ϕ80dlacZΔM15 | None | M. N. Starnbach |
| SURE | mcrA mcrCB mcrF mrr hsdR endA recB recJ F′ lacIqZΔM15 | Kan, Tet, Cm40 | Agilent Technologies |
| L. monocytogenes | |||
| EGDe | Wild-type L. monocytogenes | None | C. G. Gahan |
| ΔinlA | inlA deletion mutant derived from strain EGDe | None | C. G. Gahan |
| InlAm | Mouse-adapted L. monocytogenes; InlA S192N, Y369S | None | W.-D. Schubert |
| SD1902 | InlAm::pIMC3ery | Cm, Ery | This study |
| SD2000 | ΔinlA::pTM2 (InlAm) | None | This study |
| SD2001 | SD2000::pIMC3kan | Cm, Kan | This study |
| SD2002 | SD2000::pIMC3ery | Cm, Ery | This study |
| SD2300 | SD2000 ΔlplA1 | None | This study |
| SD2301 | SD2300::pIMC3kan | Cm, Kan | This study |
| SD2302 | SD2300::pTML1 (+lplA1) | Cm, Ery | This study |
| SD2710 | SD2000::pGJ-cGFP | Cm | This study |
| SD2800 | ΔinlA::pIMC3tet | Cm, Tet | This study |
| SD2900 | EGDe::pIMC3ery | Cm, Ery | This study |
a
Cb, carbenicillin; Cm, chloramphenicol; Ery, erythromycin; Kan, kanamycin; Tet, tetracycline.