pHHyDHODH-GFP Expression Plasmid
The pHHyDHODH-GFP expression plasmid is used to express dihydroorotate dehydrogenase (DHODH) fused to green fluorescent protein (GFP) in P. falciparum.
Highlights:
- Expression of the DHODH-GFP fusion in P. falciparum is driven by a strong HSP86 5’UTR
- Methotrexate-resistant human DHFR serves as a selectable marker for the plasmid in P. falciparum
Yeast DHODH and Aequorea GFPmut2 (1) coding sequences were inserted into an E. coli-Plasmodium falciparum shuttle plasmid (pHHMC*/3R0.5) (2), such that DHODH-GFP were fused into a single coding sequence (including a 10Ala-1Gly linker segment).
From the laboratory of Akhil B. Vaidya, PhD, Drexel University.
The pHHyDHODH-GFP expression plasmid is used to express dihydroorotate dehydrogenase (DHODH) fused to green fluorescent protein (GFP) in P. falciparum.
Highlights:
- Expression of the DHODH-GFP fusion in P. falciparum is driven by a strong HSP86 5’UTR
- Methotrexate-resistant human DHFR serves as a selectable marker for the plasmid in P. falciparum
Yeast DHODH and Aequorea GFPmut2 (1) coding sequences were inserted into an E. coli-Plasmodium falciparum shuttle plasmid (pHHMC*/3R0.5) (2), such that DHODH-GFP were fused into a single coding sequence (including a 10Ala-1Gly linker segment).
From the laboratory of Akhil B. Vaidya, PhD, Drexel University.
Specifications
| Product Type: | Plasmid |
| Gene/insert name: | DHODH |
| Accession ID: | P42212 |
| Organism: | S. cerevisiae (and P. falciparum and Homo sapiens auxiliary DNA elements) |
| Antibiotic Resistance: | Amp (E. coli) / antifolate WR99210 (P. falciparum) |
| Fusion Tag(s): | GFP/myc |
| Grow in E. coli at 37 C: | Yes |
| Selectable markers: | bla, DHFRmtxR, DHODH |
| Cloning Site 5': | BstBI |
| Cloning Site 3': | XhoI and PstI |
| Insert Size: | 942 bp |
| Vector Backbone and Size: | 6158 bp |
| High or low copy: | High |
| Promoter: | HSP86 5’UTR |
| Comments: | Note that the DHODH and GFP sequences are native sequences, i.e., not codon-optimized for either malaria or human expression. DHODH can also serve as a selectable marker by using specific inhibitors of P. falciparum DHODH or of cytochrome bc1. |
| Storage: | -20C |
| Shipped: | Room Temperature |
Provider
From the laboratory of Akhil B. Vaidya, PhD, Drexel University.
References
- Cormack B. P., et al. (1996) FACS-optimized mutants of the green fluorescent protein (GFP). Gene 173:33-8.
- O'Donnell, R. A. et al. (2002) A genetic screen for improved plasmid segregation reveals a role for Rep20 in the interaction of Plasmodium falciparum chromosomes. EMBO J 21, 1231-9.
- Painter HJ, Morrisey JM, Mather MW, Vaidya AB. (2007) Specific role of mitochondrial electron transport in blood-stage Plasmodium falciparum. Nature 446:88-91.
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