Table 1.
Plasmids and bacterial strains used in this study
| Plasmids and strains | Description or genotype | Source or reference |
|---|---|---|
| Plasmids | ||
| pMS81 | ϕBT1 attP-int-derived integration vector for the conjugal transfer of DNA from E. coli to Streptomyces spp. (Hygr) | 17 |
| pIJ10257 | 330-bp ermEp (KpnI-PstI) with ribosome binding site and multicloning site from pIJ8723 cloned into pMS81 cut with KpnI-NsiI (Hygr) | 24 |
| pIJ790 | Modified λRED recombination plasmid pKD20 | 18 |
| pGN030 | pMS81 carrying vanJ with its own promoter sequence (p-vanJ) | This study |
| pGN037 | pMS81 carrying p-vanJ with a C-terminal 6-histidine tag (C-6His tag); a stop codon (UGA) was introduced in frame immediately before the annotated start codon | This study |
| pGN038 | pMS81 carrying vanJp with a C-6His tag | This study |
| pGN034 | Short vanJ (annotated vanJ sequence) with a C-6His tag cloned in pIJ10257 under ermEp control | This study |
| pGN035 | Long vanJ (including an extra 117 bp at the 5′ end) with a C-6His tag cloned in pIJ10257 under ermEp control | This study |
| pDU001 | vanS cloned in pIJ10257 under ermEp control | Novotna and Hong (unpublished) |
| pGN091 | vanS with a C-6His tag cloned in pIJ10257 under ermEp control | This study |
| pGN024 | Long vanJ cloned in pIJ10257 under ermEp control | This study |
| pGN026 | Short staP (annotated staP sequence) cloned in pIJ10257 under ermEp control | This study |
| pGN027 | Long staP (including an extra 246 bp at the 5′ end) cloned in pIJ10257 under ermEp control | This study |
| pKV33 | SCO2255 gene cloned in pIJ10257 under ermEp control | This study |
| pKV35 | SCO7017 gene cloned in pIJ10257 under ermEp control | This study |
| pCH32 | SAV5946 gene cloned in pIJ10257 under ermEp control | This study |
| Streptomyces strains | ||
| S. coelicolor M600 | SCP1− SCP2− | 29 |
| S. coelicolor H27 | M600 + pMS81 | This study |
| S. coelicolor H2077 | M600 + pIJ10257 | This study |
| S. coelicolor H2050 | M600 + pGN024 | This study |
| S. coelicolor H357 | M600 + pKV33 | This study |
| S. coelicolor H358 | M600 + pKV35 | This study |
| S. coelicolor H359 | M600 + pCH32 | This study |
| S. coelicolor H2141 | M600 + pGN026 | This study |
| S. coelicolor H2148 | M600 + pGN027 | This study |
| S. coelicolor J3201 | ΔvanRS SCP1− SCP2− | 25 |
| S. coelicolor H102 | J3201 + pIJ10257 | This study |
| S. coelicolor H243 | J3201 + pGN024 | This study |
| S. coelioclor H2135 | J3201 + pGN026 | This study |
| S. coelicolor H2143 | J3201 + pGN027 | This study |
| S. coelicolor J3220 | ΔvanJ::apr SCP1− SCP2− | 23 |
| S. coelicolor H101 | J3220 + pMS81 | This study |
| S. coelicolor H215 | J3220 + pIJ10257 | This study |
| S. coelicolor H291 | J3220 + pGN030 | This study |
| S. coelicolor H2389 | J3220 + pGN038 | This study |
| S. coelicolor H2391 | J3220 + pGN037 | This study |
| S. coelicolor H2087 | J3220 + pGN024 | This study |
| S. coelicolor H2280 | J3220 + pGN034 | This study |
| S. coelicolor H2281 | J3220 + pGN035 | This study |
| S. coelicolor H2137 | J3220 + pGN026 | This study |
| S. coelicolor H2145 | J3220 + pGN027 | This study |
| S. coelicolor J3200 | ΔvanS SCP1− SCP2− | 25 |
| S. coelicolor H2276 | J3200 + pGN091 | This study |
| S. coelicolor J3226 | ΔvanHAX::apr SCP1− SCP2− | 23 |
| S. coelicolor H2270 | J3226 + pIJ10257 | This study |
| S. coelicolor H256 | J3226 + pGN024 | This study |
| S. coelicolor H2139 | J3226 + pGN026 | This study |
| S. coelicolor H2147 | J3226 + pGN027 | This study |
| S. coelicolor H296 | ΔSCO2255::apr SCP1− SCP2− | This study |
| S. coelicolor H347 | ΔSCO7017::apr SCP1− SCP2− | This study |
| S. toyocaensis NRRL15009 | A47934 producer; wild type | 37 |
| S. avermitilis ATCC 31267 | Avermectin producer; wild type | 44 |
| S. avermitilis H334 | ΔSAV5946::apr | This study |
| E. coli strains | ||
| ET12567(pUZ8002) | ET12567 containing helper plasmid pUZ8002 | 35 |
| BW25113(pIJ790) | BW25113 containing helper plasmid pIJ790 | 18 |