Table 1.
Strains and plasmids used in this study
| Strain or plasmid | Genotype/characteristics | Use | Reference(s) or source |
|---|---|---|---|
| Strains | |||
| Escherichia coli DH5α | fhuA2 Δ(argF-lacZ)U169 phoA glnV44 ϕ80Δ(lacZ)M15 gyrA96 recA1 relA1 endA1 thi-1 hsdR17 | Cloning | NEB |
| Escherichia coli ET12567[pUZ8002] | dam13::Tn9 dcm6 hsdM hsdR recF143 zjj201::Tn10 galK2 galT22 ara14 lacY1 xyl5 leuB6 thi1 tonA31 rpsL136 hisG4 tsx78 mtli glnV44 F − , carries plasmid pUZ8002 | Intergenic conjugation | [18, 19] |
| Streptomyces coelicolor M145 | Wild type | [17] | |
| Streptomyces coelicolor A3(2) | Wild type | [17] | |
| Streptomyces lividans TK24 | Wild type | [17] | |
| Plasmids | |||
| pCRISPomyces-2 | oriT, rpsLp (XC)-cas9 | [8] | |
| pSET152 | oriT-traJ | Source of oriT-traJ | [24] |
| pCRISPomyces-ptc | oriT, ermEp1-TheoR-cas9 | Source of TheoR | A. Rodríguez-García (unpublished) |
| pCM4.4 | oriT-traJ, ermE*p-cas9 | This work | |
| pCM4.4-Act-KO | oriT-traJ, ermE*p-cas9, spacer sequence, DNA repair template | CRISPR-Cas9-mediated deletion of actinorhodin positive transcriptional regulator | This work |
| pCM2.1 | oriT-traJ, rpsLp (XC)-cas9 | Test effect of traJ lack | This work |
| pCMU | oriT-traJ, rpsLp(XC)-TheoR-cas9 | Test effect of Cas9 levels | This work |
| pCMU-4 | oriT-traJ, D4-TheoR-cas9 | Test effect of Cas9 levels | This work |
| pCMUtuf | pCMU, tufprot | Control of Cas9 activity | This work |
| pCMU-4tuf | pCMU-4, tufprot | Control of Cas9 activity | This work |
| pCM(-cas9) | pCMU-4, Δcas9 | Control of Cas9 expression | This work |
| pCMU-4dGly | pCMU-4, glyprot, DNA repair template | CRISPR-Cas9-mediated interruption of glycerol uptake operon | This work |