Strains | Genotype | Source or reference |
---|---|---|
Escherichia coli | recA1, endA1, gyrA96, thi-1, hsdR17 (rk-, mk+), | [16] |
XL1-Blue | supE44, relA1, -, lac [F', proAB, lacIq, lacZΔM15, Tn10(Tcr)] | |
Cupriavidus necator H16 | Wildtype (PHA+) | DSMZ 428 |
Plasmids | ||
pCRII | Amp, ColE1 origin | Initrogen |
pBHR68 | PHB biosynthesis operon from C. necator in pBluescriptSK- | [17] |
pHAS | pET-14b containing NdeI/BamHI inserted phaC gene from C. necator | [18] |
pUC57-MOG | DNA fragment encoding N terminal 1–173 amino acids of MOG from mouse inserted in SmaI site of pUC57 | This study |
pUC57-IL2 | DNA fragment encoding N terminal 60–169 amino acids of IL2 from mouse inserted in SmaI site of pUC57 | This study |
pCR-phaP | pCRII containing phaP gene from C. necator | This study |
pHAS-phaP | pHAS containing the phaP gene inserted into XbaI/NdeI sites | This study |
pHAS-phaP-MOG | pHAS-phaP containing the MOG encoding DNA fragment inserted into NdeI/BamHI sites | This study |
pHAS-phaP-MOG | pHAS-phaP containing the IL2 encoding DNA fragment inserted into NdeI/BamHI sites | This study |
pUC57-phaP-MOG | DNA fragment encoding PhaP inserted into XbaI and NdeI site of plasmid pUC57-MOG | This study |
pUC57-phaP-IL2 | DNA fragment encoding PhaP inserted into XbaI and NdeI site of plasmid pUC57-IL2 | This study |
pBHR68-phaP-MOG | DNA fragment encoding PhaP-MOG fusion protein subcloned from pUC57-phaP-MOG via XbaI and BamHI into pBHR68 | This study |
pBHR68-phaP-IL2 | DNA fragment encoding PhaP-IL2 fusion protein subcloned from pUC57-phaP-IL2 via XbaI and BamHI into pBHR68 | This study |
Oligonucleotides | ||
phaP-XbaI-NdeI | 5'-aaaaatctagaaaaaggagatatacatatgatcctc-accccggaacaag-3' | This study |
phaP-XbaI | 5'-aaaaatctagaaaaaggagatatacgtatgatcctc-accccggaacaag-3' | This study |
phaP-NdeI | 5'-aaaaacatatggtggtgatggtgatgcgagccgcgt-ttatcatcatcatcggcagccgtcgtcttc-3' | This study |