TABLE 1.
Strains and vectors used in this study
| Strain, plasmid, or cosmid | Genotype and characteristics | Reference or source |
|---|---|---|
| Strains | ||
| Streptomyces coelicolor A3(2) | ||
| M145 (wild type) | SCP1− SCP2− | 15 |
| NM3 (Δnar-2) | M145 ΔSCO0216-ΔSCO0219::aac(3)IV (deletion of 6,531 bp removing narG2H2J2I2) | 12 |
| NM24 (Δnar-1) | M145 ΔSCO6535-ΔSCO6532::aadA (deletion of 6,209 bp removing narG1H1J1I1) | 13 |
| NM27 (Δnar-3) | M145 ΔSCO4947-ΔSCO4950::aadA (deletion of 6,497 bp removing narG3H3J3I3) | 13 |
| NM29 (Δnar-1 Δnar-2) | NM4 ΔSCO6535-ΔSCO6532::aadA (deletion of 6,209 bp removing narG3H3J3I3) | 13 |
| NM59 (Δnar-1 Δnar-3) | NM39 ΔSCO6535-ΔSCO6532::aadA (deletion of 6,209 bp removing narG3H3J3I3) | 13 |
| NM68 (Δnar-2 Δnar-3) | NM3 ΔSCO4947-ΔSCO4950::aadA (deletion of 6,497 bp removing narG3H3J3I3) | 12 |
| NM92 (Δnar-1 Δnar-2 Δnar-3) | NM90 ΔSCO6532-ΔSCO6535::aadA (deletion of 6,209 bp removing narG1H1J1I1) | 12 |
| COE285 | M145 SCO0213::Tn5062 (ΔnarK2) | This study |
| COE404 | COE285 with integrative vector pMSnarK2 (SCO0213+) | This study |
| COE301 | M145 SCO2959::Tn5062 (ΔnarK1) | This study |
| Escherichia coli | ||
| DH5α | F− ϕ80dlacZΔM15 endA recA hsdR(rK− mK−) supE thi-1 gyrA relA Δ(lacZYA-argF)U169 | Laboratory stock |
| ET12567(pUZ8002) | Δdam dcm with trans-mobilizing plasmid pUZ8002 | 16 |
| JTU007(pUZ8002) | Δdam Δdcm with trans-mobilizing plasmid pUZ8002 | 17 |
| Rosetta(DE3)/pLysS | F− ompT hsdSB(rB− mB−) gal dcm λ(DE3 [lacI lacUV5-T7 gene 1 ind1 Sam7 nin5])/pLysSRARE (CLMr) | Novagen |
| Plasmids and cosmids | ||
| SCJ12.2.03_04081112EO | Cosmid SCJ12 disrupted in SCO0203 with Tn5062 | 18 |
| SCJ12.1.F10_040807014A | Cosmid SCJ12 disrupted in SCO0213 with Tn5062 | 18 |
| pMS82 | ΦBT1 attP-int-derived integration vector for conjugal transfer of DNA from E. coli to Streptomyces (Hygr) | 19 |
| pMSnarK2 | pMS82 SCO0213 (with 236-bp upstream and 227-bp downstream sequences) | This study |
| pET30a | (+) Expression plasmid, Kanr | Novagen |
| pET30-narG2 | Like pET30a(+) but narG2 positive | This study |