TABLE 1.
Plasmids and strains used in this study
| Plasmid or strain | Relevant characteristic(s) | Source or reference |
|---|---|---|
| Plasmid | ||
| pAE5 | Encodes B. cereus 569 plcR under control of B. anthracis protective antigen gene (pagA) promoter; Apr in E. coli; Kmr in B. anthracis | (Pomerantsev et al., 2003) |
| pAE51 | pAE5 with stop codon in plcR truncating six C-terminal amino acids | This work |
| pUTE29-plcR-papR | B. cereus 569 plcR-papR DNA fragment inserted into pUTE29; Apr in E. coli; Tcr in B. anthracis | (Pomerantsev et al., 2004) |
| pFP12 | pUTE29-plcR fusion to papR | (Pomerantsev et al., 2004) |
| pUTE29-papR | B. cereus 569 papR DNA fragment inserted into pUTE29 | (Pomerantsev et al., 2004) |
| p102 | pUTE29-nprB-plcR-papR | This work |
| p182 | pUTE29-nprB-plcR fusion to papR | This work |
| pUTE29-nprB | P182 without BstZ17I-PstI fragment encoding fused PlcR-PapR | This work |
| pJRS312 | pUC18 carrying an Ω element with spectinomycin resistance marker (Ω-sp) | J.R. Scott |
| p102Sp | p102 with Ω-sp cassette flanked by two directly repeated loxP and inserted into BglII-site of nprB | This work |
| p182Sp | p182 with Ω-sp cassette flanked by two directly repeated loxP and inserted into BglII-site of nprB | This work |
| pHY304 | Contains strongly temperature-sensitive replicon for both E. coli and gram-positive bacteria; Emr in both E. coli and B. anthracis | (Pritzlaff et al., 2001) |
| pHU304 | Large PvuII-fragment of pUC18 combined with large Ecl136II-fragment of pHY304 | This work |
| pHU102 | PvuII fragment of p102 containing nprB- plcR-papR gene cluster inserted into BstZ17I-site of pHU304 | This work |
| pHU102Sp | PvuII fragment of p102Sp containing nprB∷(loxP-Ω-sp-loxP) - plcR-papR gene cluster inserted into BstZ17I-site of pHU304 | This work |
| pCrePA | pHY304 with Cre recombinase gene under control of pagA promoter | (Pomerantsev et al., 2006) |
| pDG148-Stu | Gram-Positive-E. coli shuttle vector featuring ligation-independent cloning and inducible expression; Apr in E. coli; Kmr in B. anthracis | (Joseph et al., 2001) |
| pB148 | pDG148-Stu carrying B. cereus 569 nprB gene under control of Pspac promoter | This work |
| pB148Lac- | pB148 without lacI repressor gene | This work |
| Strain | ||
| B. cereus | ||
| 569 | Wild strain, produces extracellular PC-PLC | (Reddy et al., 1987) |
| 541 | 569; intact plcR lacks a promoter; Emr; does not produce extracellular PC-PLC | (Pomerantsev et al., 2003) |
| 537 | 569; nprB knockout, Spr; nprB contains Ω-sp cassette flanked by two directly repeated loxP; demonstrates very low PC-PLC activity | This work |
| 536 | 569; nprB knockout containing one loxP site; demonstrates very low PC-PLC activity | This work |
| 535 | 569; papR replaced by loxP site; demonstrates very low PC-PLC activity | This work |
| B. anthracis | ||
| SdT | Sterne 34F2 cured of pXO1; therefore pXO1- pXO2- | (Ivins et al., 1986) |
| SdT2 | SdT electroporated with pAE5; Kmr; produces intracellular PlcR; does not produce a halo when grown on LB agar with lecithin | (Pomerantsev et al., 2003) |
| SdT251 | SdT electroporated with pAE51; Kmr; produces intracellular PlcR with truncated six C-terminal amino acids; does not produce a halo when grown on LB agar with lecithin | This work |
| SdT12 | SdT electroporated with pFP12; Tcr; produces intracellular PlcR-PapR fused protein; demonstrates does not produce a halo when grown on LB agar with lecithin | (Pomerantsev et al., 2004) |
| Ames 33 | pXO1- pXO2- Ames derivative strain | (Pomerantsev et al., 2003) |
| BH450 | Ames 33 ba_0599 protease knockout, spo0A knockout | (Pomerantsev et al., 2006) |