Table 2 Strains, genes, and plasmids used in this study
From: Development of a thiostrepton-free system for stable production of PLD in Streptomyces lividans SBT5
Strains/genes/plasmids | Characteristics | Reference |
|---|---|---|
Strains | ||
E. coli DH5α | General cloning host | [53] |
E. coli ET12567/pUZ8002 | Methylation-deficient strain, for E. coli-Streptomyces interspecies conjugation | [54] |
S. antibioticus | Wild-type strain | [9] |
S. lividans SBT5 | Expression host. S. lividans TK24 △act△redKL△cdaPS3-SLI3600::afsRScla | [24] |
Genes | ||
kasOp* | Constitutive strong promoter for gene overexpression in Streptomyces (GenBank: MN306530) | [30] |
sigcin | Stv. cinnamoneum pld (GenBank: BAA75216) derived signal peptide | [39] |
mpld | S. antibioticus pld (GenBank: BAA03913) was codon-optimized, G215S mutation | [38] |
pld* | kasOp*-sigcin-mpld | |
sti | Streptomyces strong incompatibility region (GenBank: AY667410) | [36] |
sta | The sequence from pIJ101 between the PstI site at position 31 and the SstII site at position 33 | [31] |
rep3 | Derived from the Streptomyces vector pIJ6021 (GenBank: AJ414669). sta, repP, orf56 | [33] |
Plasmids | ||
pIJ101 | Multi-copy broad host-range Streptomyces plasmid (GenBank: M21778) from S. lividans ISP 5434 | [31] |
pSET152 | φC31 based streptomyces integrative vector, Apr r | [42] |
pJTU1278 | Streptomyces autonomously replicating vector, oriT (RK2), pIJ101 ori, ColE1 ori, Ampr, Tsr r | [35] |
pOJ260 | Vector for gene disruption and transplacement, oriT (RK4), pUC18 ori, Aprr | [42] |
pUC57 | E. coli cloning vector | |
pUC57-p | Artificially synthesized sequence pld* EcoRV cloned into vector pUC57, Ampr | This work |
pUC57-s | Artificially synthesized sequence sti EcoRV cloned into vector pUC57, Ampr | This work |
pUC57-r | Artificially synthesized sequence rep3 EcoRV cloned into vector pUC57, Ampr | This work |
pOJ260-rep1 | Fragment rep from pJTU1278 SpeI cloned into vector pOJ260. repP, orf56, Aprr | This work |
pOJ260-rep2 | Fragment sti from pUC57-s KpnI/SpeI cloned into vector pOJ260-rep1. sti, repP, orf56, Aprr | This work |
pOJ260-rep3 | Fragment rep3 from pUC57-r SpeI cloned into vector pOJ260. sta, repP, orf56, Aprr | This work |
pSET152-pld* | Fragment pld* from pUC57-p XbaI cloned into vector pSET152, Aprr | This work |
pOJ260-repn-pld* | Fragment pld* XbaI cloned into vector pOJ260-repn, Aprr. repn: rep1, rep2, rep3 | This work |