Table 1.
Bacterial strains and plasmids
| Strains | Genotype/relevant phenotype | Origin |
|---|---|---|
| E. faecalis | ||
| OG1RF | Clinical isolate | (Murray et al., 1993) |
| V583 | Clinical isolate, TIGR sequence strain; VnR | (Thomas et al., 2008) |
| VT01 | V583ΔgelE; SprE+ | (Thomas et al., 2008) |
| VT02 | V583ΔsprE; GelE+ | (Thomas et al., 2008) |
| VT03 | V583ΔgelE-sprE | (Thomas et al., 2008) |
| VT12 | VT03 (pMV158GFP); GFP+ TetR | (Thomas et al., 2008) |
| VT13 | V583::fsrA (pVT31); CmR, SpcR | This study |
| VT14 | V583 (pVT30); SpcR | This study |
| VT15 | V583 (pVT31); GFP+ SpcR | This study |
| VT16 | VT01 (pVT31); GFP+ SpcR | This study |
| VT17 | VT02 (pVT31); GFP+ SpcR | This study |
| VT18 | VT03 (pVT31); GFP+ SpcR | This study |
| VT19 | V583:: atlA; CmR | This study |
| VT20 | OG1RFΔgelE; SprE+ | This study |
| VT21 | OG1RF:: atlA; CmR | This study |
| VT22 | VT03::atlA; CmR | This study |
| VT23 | VT12::atlA (pMV158GFP); GFP+, CmR, TetR | This study |
| VT24 | VT03 (pVT08); SprE+ | This study |
| KS10 | V583ΔfsrA | Gorman, M. and Hancock, L.E., (unpublished) |
| E. coli | ||
| Electro Ten Blue | General plasmid maintenance strain | Stratagene |
| BL21 (DE3) | Protein expression strain | Stratagene |
| VT25 | BL21(DE3) (pVT21; AtlA+ * AmpR | This study |
| Plasmids | ||
| pAT28 | Broad host range shuttle vector; SpcR | (Trieu-Cuot et al., 1990) |
| p3CAT | p3TET derivative (Hancock and Perego, 2004), suicide vector; CmR, used for targeted gene inactivation | Hancock, LE (unpublished) |
| pMV158GFP | Gram-positive replicative vector expressing a GFP reporter | (Nieto and Espinosa, 2003) |
| pML115 | p3CAT derivative carrying an internal fragment of fsrA; generates fsrA::cm mutation in E. faecalis | Hancock, L.E (unpublished) |
| pVI01 | Low copy gelE promoter-gfp fusion reporter construct | Iyer, V and Hancock, L.E (unpublished) |
| pVT08 | pAT28 derivative carrying a 2123-bpEcoRI/BamHI fragment (native gelE promoter and full-length sprE); leads to the over expression of SprE | (Thomas et al., 2008) |
| pVT10 | p3CAT derivative carrying a 1006-bp EcoR1/BamH1 internal fragment of atlA; generates atlA::cm mutation in E. faecalis | This study |
| pVT21 | pET21b derivative carrying a 2056-bp Nde1/Xho1 atlA fragment devoid of signal peptide; used for the expression of AtlA in E. coli | This study |
| pVT30 | pAT28 derivative carrying a 759-bp SalI/SalI gfp fragment; promoterless gfp | This study |
| pVT31 | pAT28 derivative carrying a gelE promoter-gfp fusion; reporter construct that allows discrimination of predators within E. faecalis population | This study |
R
Resistant
Vn, vancomycin; Cm, chloramphenicol; Spc, spectinomycin; Tet, tetracycline; Amp, ampicllin
*
AtlA devoid of signal peptide (53 amino acid residues from the N-terminus)