TABLE 1.
Bacterial strains and plasmids used in this study
| Strain or plasmid | Relevant characteristic(s) | Source or reference |
|---|---|---|
| Strains | ||
| Escherichia coli DH5α | F−endA1 hsdR17(rK− mK+) supE44 thi-1 λ−gyrA96 relA1 Δ(lacZYA argF)U169 (φ80 lacZ ΔM15) recA1 | Roche Diagnostics |
| Clostridium cellulolyticum | ||
| ATCC 35319 | Wild type | 24 |
| cipCMut1 | cipC::ISCce1::ISCce2 | 16 |
| cipCMut1 (pSOSzero) | cipCMut1 pSOSzero transformant | This study |
| cipCMut1 (pSOScipC) | cipCMut1 pSOScipC transformant | 16 |
| Plasmids | ||
| pSOSzero | pSOS95 derivative with entire expression cassette deleted | 23 |
| pSOScipC | pSOS952 derivative expression vector bearing cipC gene | 16 |
| pZ41 | pUC18 derivative carrying 6.3-kb PvuII fragment bearing cel9G gene | 1 |
| pLM26 | pUC13 derivative carrying 5-kb HindIII/Sau3AI fragment bearing cel8C | 1 |
| pETFc | E. coli expression vector bearing cel48F gene | 26 |
| pSPT18 | Cloning vector; Apr | Roche Diagnostics |
| pSPTcipC | pSPT18 derivative carrying 1.1-kb HindIII/BamHI fragment of cipC | 16 |
| pSPTcel48F | pSPT18 derivative carrying 0.9-kb BamHI/Asp718 fragment of cel48F | This study |
| pSPTcel8C | pSPT18 derivative carrying 0.9-kb PvuII fragment of cel8C | This study |
| pSPTcel9G | pSPT18 derivative carrying 0.75-kb HindIII/BglII fragment of cel9G | This study |
| pSPTcel9E | pSPT18 derivative carrying 0.9-kb EcoRI-digested PCRa fragment of cel9E | This study |
| pSPTcel9H | pSPT18 derivative carrying HincII/EcoRV-digested PCRa fragment of cel9H | This study |
| pSPTcel9J | pSPT18 derivative carrying PstI/PvuII-digested PCRa fragment of cel9J | This study |
| pSPTrgl11Y | pSPT18 derivative carrying SspI/HincII-digested PCRa fragment of rgl11Y | This study |
| pGEM-T Easy | Cloning vector; Apr | Promega |
| pGEMorfX | pGEM-T Easy derivative carrying PCRa fragment of orfX | This study |
| pGEMman5K | pGEM-T Easy derivative carrying BspEI/NcoI-digested PCRa fragment of man5K | This study |
| pGEMcel9M | pGEM-T Easy derivative carrying PCRa fragment of cel9M | This study |
| pGEMcel5N | pGEM-T Easy derivative carrying PCRa fragment of cel5N | This study |
| pGEM16S | pGEM-T Easy derivative carrying PCRa fragment of 16S gene | This study |
a
The primers used to synthesize the PCR fragments are listed in Table 2.