TABLE 1.
Strains and plasmids used in this study
| Strain or plasmid | Description | Reference no. or source |
|---|---|---|
| C. difficile strains | ||
| R20291 | Wild type, ribotype 027 | 55 |
| CRG2359 | R20291 ΔpyrE | 35 |
| RS19 | CRG2359-restored pyrE | 38 |
| MB03 | ΔspoVAD strain | This study |
| MB04 | ΔspoVAE strain | This study |
| Other strains | ||
| E. coli DH5α | Cloning strain | 51 |
| E. coli HB101 pRK24 | Conjugal donor strain for C. difficile | 56 |
| Plasmids | ||
| pMTL-YN4 | Backbone used to make deletions in R20291 by allelic exchange | 35 |
| pMTL84151 | Backbone used to make complementing plasmids and spore luciferase interaction plasmids | 57 |
| pMB02 | To create ΔspoVAD deletion | This study |
| pMB04 | To create ΔspoVAE deletion | This study |
| pMB34 | spoVAD-complementing plasmid | This study |
| pMB35 | spoVAE-complementing plasmid | This study |
| pAP118 | Plasmid encoding aTC-inducible HupA-SmBit and HupA-LgBit fusions | 43 |
| pAF256 | Plasmid encoding an aTC-inducible HupA-SmBit and LgBit | 43 |
| pAF257 | Plasmid encoding an aTC-inducible SmBit and HupA-LgBit | 43 |
| pAF259 | Plasmid encoding an aTC-inducible BitlucOpt | 43 |
| pMB50 | Luciferase SpoVAC/SmBit-LgBit | This study |
| pMB51 | Luciferase SpoVAD/SmBit-LgBit | This study |
| pMB52 | Luciferase SpoVAE/SmBit-LgBit | This study |
| pMB53 | Luciferase SmBit-SpoVAC/LgBit | This study |
| pMB54 | Luciferase SmBit-SpoVAD/LgBit | This study |
| pMB55 | Luciferase SmBit-SpoVAE/LgBit | This study |
| pMB62 | Luciferase SpoVAC/SmBit-SpoVAC/LgBit | This study |
| pMB45 | Luciferase SpoVAC/SmBit-SpoVAE/LgBit | This study |
| pMB46 | Luciferase SpoVAD/SmBit-SpoVAC/LgBit | This study |
| pMB47 | Luciferase SpoVAD/SmBit-SpoVAD/LgBit | This study |
| pMB48 | Luciferase SpoVAD/SmBit-SpoVAE/LgBit | This study |
| pMB49 | Luciferase SpoVAE/SmBit-SpoVAC/LgBit | This study |
| pMB57 | Luciferase SpoVAE/SmBit-SpoVAE/LgBit | This study |
| pMB60 | Luciferase SpoVAC/SmBit-SpoVAD/LgBit | This study |
| pMB61 | Luciferase SmBit-LgBit | This study |
| pMB72 | SpoVAC-SmBit/SpoVAE-LgBit luciferase spore complementation | This study |
| pMB73 | SpoVAD-SmBit/SpoVAC-LgBit luciferase spore complementation | This study |
| pMB75 | SpoVAD-SmBit/SpoVAE-LgBit luciferase spore complementation | This study |
| pMB79 | HupA-SmBit/HupA-LgBit luciferase spore complementation | This study |
| pMB81 | BitLuc luciferase spore complementation | This study |
| pMB82 | SpoVAC-SmBit-SpoVAC/SpoVAD-LgBit; small luciferase reporter inserted mid-AC gene, ADLg partner | This study |
| pMB83 | SpoVAD-SmBit/SpoVAC-LgBit-SpoVAC; large luciferase reporter inserted mid-AC gene, ADSm partner | This study |
| pMB84 | SpoVAC-SmBit-SpoVAC/SpoVAE-LgBit; small luciferase reporter inserted mid-AC gene, AELg partner | This study |
| pMB85 | SpoVE-SmBit/SpoVAC-LgBit-SpoVAC; large luciferase reporter inserted mid-AC gene, AESm partner | This study |
| pMB87 | Sm-Lg luciferase-spore interaction plasmid, negative control | This study |
| pMB88 | SpoVAD-Sm/SpoVAE-Lg-SpoVAE; large luciferase reporter inserted mid-AE gene, ADSm partner | This study |
| pMB89 | SpoVAE-Sm-SpoVAE/SpoVAD-Lg; small luciferase reporter inserted mid-AE gene, ADLg partner | This study |