Table 1.
Bacterial strains and plasmids used in this study
| Strain or plasmid | Relevant descriptiona | Reference or source |
|---|---|---|
| Strains | ||
| G. kaustophilus | ||
| HTA426 | Wild type | JCM 12893 |
| MK54 | ΔpyrF | This study (Fig. 3) |
| MK54c | Mutant MK54 that integrates pGAM46 at GK0707 locus | This study |
| MK72 | ΔpyrF ΔpyrR | This study (Fig. 5) |
| MK72c | Mutant MK72 that integrates pGAM46 at GK0707 locus | This study |
| MK95 | Derivative of mutant MK72; GK0707::PsigA-bgaB | This study (Fig. 6) |
| MK96 | Derivative of mutant MK72; GK0707::bgaB | This study (Fig. 6) |
| MK109 | Derivative of mutant MK72; GK0707::PsigA-amyE | This study (Fig. 6) |
| MK304 | Derivative of mutant MK72; GK0707::amyE | This study (Fig. 6) |
| G. stearothermophilus | ||
| IAM11011 | Source of bgaB | ATCC 8005 |
| CU21 | Source of amyE | ATCC 12980 |
| E. coli BR408 | Donor strain for conjugative plasmid transfer | 28 |
| Plasmids | ||
| pUC19 | Cloning vector; Ampr | TaKaRa Bio (Fig. 2) |
| pCR4Blunt-TOPO | Cloning vector; Ampr Kmr | Invitrogen |
| pCR4B-TK101 | pCR4Blunt-TOPO carrying TK101 marker | This study |
| pCR4B-pyrF | pCR4Blunt-TOPO carrying pyrF marker | This study |
| pGKE11T | Derivative of pUC19; oriT Ampr | This study (Fig. 2) |
| pTK19 | Derivative of pGKE11T, oriT, Ampr, TK101 marker | This study (Fig. 2) |
| pΔpyrF | pTK19 carrying ΔpyrF fragment | This study (Fig. 2) |
| pΔpyrR | pTK19 carrying ΔpyrR fragment | This study (Fig. 2) |
| pGAM46 | Derivative of pGKE11T, GK0707 fragments, oriT, Ampr, pyrF marker | This study (Fig. 2) |
| pGAM46-bgaB | pGAM46 carrying bgaB between SphI and BamHI sites | This study |
| pGAM46-amyE | pGAM46 carrying amyE between SphI and BamHI sites | This study |
| pGAM47 | pGAM46 carrying PsigA between HindIII and SphI sites | This study |
| pGAM47-bgaB | pGAM47 carrying bgaB between SphI and BamHI sites | This study |
| pGAM47-amyE | pGAM47 carrying amyE between SphI and BamHI sites | This study |
a
Ampr and Kmr are genes coding for resistance to ampicillin and kanamycin, respectively. oriT is the conjugative transfer origin from pRK2013 (9). PsigA is the sigA promoter from G. kaustophilus HTA426. The TK101 marker represents the TK101 gene encoding thermostable kanamycin nucleotidyltransferase (18) under the control of the sigA promoter. The pyrF marker is the pyrF gene from G. kaustophilus HTA426 under the control of the sigA promoter.