TABLE 1.
Bacterial strains, plasmids, and antibiotic markers
| Strain or plasmid | Characteristic(s) | Antibiotica | Source or reference |
|---|---|---|---|
| Strains | |||
| C. tepidum | |||
| WT2321 | Wild type | None | 59 |
| CT0117::TnOGm | TnOGm insertion in CT0117 | Gm | This study |
| CT0876::TnOGm | TnOGm insertion in CT0876 | Gm | This study |
| CT1087::TnOGm | TnOGm insertion in CT1087 | Gm | This study |
| CT0117::TnOGm, CT1087::TnEm | TnOGm and TnEm insertions in CT0117 and CT1087, respectively | Gm, Em | This study |
| CT1087-His6::aacC1 | Encodes CT1087 protein with C-terminal His6 extension, followed by the insertion of an aacC1 gene downstream of the CT1087 coding sequence | Gm | This study |
| E. coli strains | |||
| TOP10 | General cloning strain; F−mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 recA1 araΔ139 Δ(ara-leu)7697 galU galK rpsL (Strr) endA1 nupG | None | Invitrogen |
| BL21λDE3/pLysS | Recombinant protein expression strain; ompT hsdSB(rB− mB−) gal dcm | Cm | Novagen |
| Plasmids | |||
| pCR2.1 | Cloning vector | Km | Invitrogen |
| pET16b | T7 expression vector, N-terminal His10 tag | Car | Novagen |
| pET28b | T7 expression vector, N- and C-terminal His6 tags | Km | Novagen |
| pET28b-CT0117 | CT0117 cloned into EcoRI-HindII sites | Km, Cm | This study |
| pET16b-CT0876 | CT0876 cloned into NdeI-BamHI sites | Car, Cm | This study |
| pET16b-CT1087 | CT1087 cloned into NdeI-BamHI sites | Car, Cm | This study |
a
Gm, gentamicin (4 μg ml−1 for liquid medium and 8 μg ml−1 for plates); Em, erythromycin (for C. tepidum, 5 μg ml−1 for liquid medium and 10 μg ml−1 for plates; for E. coli, 20 μg ml−1, used to construct a plasmid to create CT0117::TnOGm and CT1087::TnEm); Cm, chloramphenicol (20 μg ml−1); Km, kanamycin (50 μg ml−1); Car, carbenicillin (50 μg ml−1).