TABLE 1.
Bacterial strains and plasmids used in this study
| Strain or plasmid | Relevant characteristics | Reference or source |
|---|---|---|
| E. coli strain DH5α | supE44 hsdR17 recA1 endA1 gyrA96 thi-1 relA1 | 12 |
| N. meningitidis strains | ||
| MC58 | Clinical isolate; sequenced strain | 29 |
| wtA (MC-PcrgA1) | crgA promoter region from position −63 to position 179, fused to lacZ, is inserted between ORFs NMB1074 and NMB1075; Eryr | This study |
| Δ35 (MC-PcrgA2) | crgA promoter region from position −15 to position 179, fused to lacZ, is inserted between ORFs NMB1074 and NMB1075; Eryr | This study |
| Δ10 (MC-PcrgA3) | crgA promoter region from position −6 to position 179, fused to lacZ, is inserted between ORFs NMB1074 and NMB1075; Eryr | This study |
| ΔP (MC-PcrgA4) | crgA promoter region from position 44 to position 179, fused to lacZ, is inserted between ORFs NMB1074 and NMB1075; Eryr | This study |
| ΔR (MC-PcrgA5) | crgA promoter region from position −63 to position 43, fused to lacZ, is inserted between ORFs NMB1074 and NMB1075; Eryr | This study |
| ΔRCrgA (MC-C−PcrgA5) | CrgA null mutant, derivative of ΔR (MC-PcrgA5); Eryr Kmr | This study |
| ΔRCrgA-C (MC-C−CindPcrgA5) | CrgA complemented mutant, derivative of ΔRCrgA (MC-C−PcrgA5) containing the crgA gene under the control of the Ptac promoter and lacI repressor located between ORFs NMB1428 and NMB1429; Eryr Kmr Cmr | This study |
| Plasmids | ||
| pGem3Z | Cloning vector, Ampr | Promega |
| pSL1190 | Cloning vector, Ampr | Pharmacia |
| pCMVβ | Plasmid containing the lacZ gene of E. coli | Clontech |
| pAT110 | Plasmid containing the ermAM erythromycin resistance gene | 30 |
| pDT2548 | Plasmid containing the chloramphenicol resistance cassette from Campylobacter coli | 31 |
| pILL600 | Plasmid containing the kanamycin cassette from Campylobacter coli | 16 |
| pMMB206 | Plasmid containing the Ptac promoter and the lacI gene | 19 |
| pSL-Fla-Ery | Plasmid consisting of a promoterless lacZ gene and the ermAM erythromycin resistance gene flanked by upstream and downstream regions for allelic replacement at a chromosomal location between ORFs NMB1074 and NMB1075 | This study |
| pSL-Pcrg1 | Derivative of pSL-Fla-Ery containing the full crgA promoter region from position −63 to position 179 fused to the lacZ gene | This study |
| pSL-Pcrg2 | Derivative of pSL-Fla-Ery containing the crgA promoter region from position −15 to position 179 fused to the lacZ gene | This study |
| pSL-Pcrg3 | Derivative of pSL-Fla-Ery containing the crgA promoter region from position −6 to position 179 fused to the lacZ gene | This study |
| pSL-Pcrg4 | Derivative of pSL-Fla-Ery containing the crgA promoter region from position 44 to position 179 fused to the lacZ gene | This study |
| pSL-Pcrg5 | Derivative of pSL-Fla-Ery containing the crgA promoter region from position −63 to position 43 fused to the lacZ gene | This study |
| pG3cr:Km | Plasmid for knockout of crgA gene containing the upstream and downstream regions of the crgA locus flanking a kanamycin resistance cassette | This study |
| pSLComCmr | Plasmid consisting of the chloramphenicol resistance gene flanked by upstream and downstream regions for allelic replacement at a chromosomal location between ORFs NMB1428 and NMB1429 | This study |
| pPindcrgA | Plasmid for complementation of the CrgA null mutant, derivative of pSLComCmr containing a copy of the crgA gene under the control of the Ptac promoter and the lacI repressor | This study |