TABLE 1.
Bacterial strains and plasmids used
| Strain or plasmid | Characteristics | Reference or source |
|---|---|---|
| Strains | ||
| E. coli | ||
| JM109 | recA1 supE44 endA1 hsdR17 gyrA96 relA1 thi Δ(lac-proAB) | 39 |
| S17-1 | thi1 proA hsdR17 recA1 tra (RP4) | 27 |
| SJ16 | F−panD2 zad-220::Tn10 λ−216 relA1 spoT1 metB1 λr Tcr | 10 |
| R. eutropha | ||
| HF39 | Parent strain Smr | 29 |
| C3 | pntAA::Tn5 derived from R. eutropha HF39, β-alanine auxotroph | This study |
| M30 | pntAA::Tn5 derived from R. eutropha HF39, β-alanine auxotroph | This study |
| O22 | panD::Tn5 derived from R. eutropha HF39, β-alanine auxotroph | This study |
| Q20 | panD::Tn5 derived from R. eutropha HF39, β-alanine auxotroph | This study |
| Plasmids | ||
| pVK100 | Cosmid, Tcr, Kmr | 14 |
| pBluescript SK− | Apr, ColE1 | Stratagene |
| pBluescript KS− | Apr, ColE1 | Stratagene |
| pBBR1MCS-3 | Tcr, RK2, Mob+, Tra− | 15 |
| pBHR68 | Derivative of pBluescript SK− containing the 5.2-kb SmaI/EcoRI fragment comprising the PHA operon from R. eutropha | 28 |
| pBHR68(S260A) | Like pBHR68 but containing site-specific mutant of phaC gene (serine at position 260 changed to alanine) | This study |
| pBHR68(S546I) | Like pBHR68 but containing site-specific mutant of phaC gene (serine at position 546 was changed to isoleucine) | This study |
| pVK100-C20 | Derivative of pVK100 containing a multiple 33.5-kb EcoRI fragment from R. eutropha | This study |
| pSKE15 | Derivative of pBluescript SK− containing a 15-kb EcoRI fragment subcloned from pVK100-C20 | This study |
| pKSKX0.76 | Derivative of pBluescript KS− containing a 0.76-kb PCR-amplified KpnI/XhoI fragment comprising the coding region of panD from R. eutropha | This study |
| pBBR1MCS-3KX0.76 | Derivative of pBBR1MCS-3 containing a 0.76-kb PCR-amplified KpnI/XhoI fragment comprising the coding region of panD from R. eutropha | This study |