Last data update: 24 January 2024 16:39 CET
Plasmid name: pDESTMB14-noGFP-unc54-pRPS-0-GAS-1-roGFP2-ORP-1 (LMBP 9708)
| New search | Print data sheet |
| Price category: | Cat. 3 (cf. price list) |
| Status: | GeneCorner non-core plasmid |
| Depositor's sequence: | not available |
| Sequence analysis results Genecorner: |
NGS: b05-gc-dec2018.fasta |
| Cloned DNA: |
Caenorhabditis elegans general anaesthetic sensitivity abnormal cDNA (gas-1) Aequorea victoria green fluorescent protein DNA (GFP); reduction-oxidation sensitive variant (roGFP), cytosol variant (roGFP2) Glutathion peroxidase 3 (GPX3, ORP1) |
| Promoter: | Caenorhabditis elegans 40S ribosomal protein SA promoter (rps-0) Escherichia coli lac operon promoter Phage T3 promoter Phage T7 gene 10 promoter (T7g10) |
| Ribosome binding site: |
- |
| Terminator: | Caenorhabditis elegans unc-54 terminator (unc-54) |
| Selection marker: | Ampicillin (amp) Caenorhabditis elegans unc-119 (unc-119); auxotrophic |
| Replicon: | Escherichia coli plasmid pMB1 origin Phage f1 origin |
| Host range: | Escherichia coli Caenorhabditis elegans; unc-119(-) |
| Parental clone: | pENTRYL4R1-pRPS-0; pDestMB14 |
| Further information: | This Gateway Expression vector was created by removing the GFP coding sequence from pDestMB14 and cloning the gas-1-roGFP2-GPX3 cassette into this destination vector, behind the C. elegans rps-0 promoter obtained from pENTRYL4R1-pRPS-0. The reduction-oxidation sensitive green fluorescent protein (roGFP) is a redox sensitive biosensor. Two cysteines were introduced into the beta barrel structure of the GFP. The oxidation state (reduced dithiol or the oxidized disulfide) of the engineered thiols determines the fluorescence properties of the sensor. RoGFP2 preferentially interacts with glutaredoxins and therefore reports the cellular glutathione redox potential. The specificity of roGFP2 for glutathione is further increased by linking it to GRX3. By expressing the roGFP2-GRX3 fusion sensors in the organism of interest and/or targeting the protein to a cellular compartment, it is possible to measure the glutathione redox potential in a specific cellular compartment in real-time and therefore provides major advantages compared to other invasive static methods e.g. HPLC. The nucleotide sequence of the C. elegans inserts can be retrieved from the Genbank accession numbers NC_003284.9, NC_003281.10 and NC_003279.8. Other name of the plasmid is pDESTM14-NOGFP+UNC54-prpso-Gas1-roGFP2-Orp1 in DH5a (E. Coli). |
| EMBL Accession number: | NC_003284.9, view at EMBL, GenBank, DDBJ NC_003281.10, view at EMBL, GenBank, DDBJ NC_003279.8, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 09/12/2015 |
| Authenticity test: | This plasmid has been fully sequenced but the NGS sequence data still need to be implemented. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by C. Vlaeminck(1) and Prof. Dr B. Braeckman(1). (1) Research Group for aging physiology and molecular evolution, Department of Biology, Ghent University, Ghent |
| Plasmid reference: | - |
| Restricted distribution: | - BCCM MTA |
| Distributed as: | active culture or plasmid DNA |
| Host for distribution: | Escherichia coli K12 DH5α |
| Host reference: | Focus 8 (1986), 9 |
| Related host reference: | Woodcock et al., Nucleic Acids Res. 17 (1989), 3469-3478 [PMID: 2657660] Rodriguez-Quinones et al., Focus 15 (1993), 110-112 Hanahan, J. Mol. Biol. 166 (1983), 557-580 [PMID: 6345791] [DOI: 10.1016/s0022-2836(83)80284-8] Hanahan, in 'DNA Cloning: A Practical Approach Volume I', IRL Press, Oxford (1985), 109-135 [ISSN/ISBN: 0947946187] Grant et al., Proc. Natl. Acad. Sci. U.S.A. 87 (1990), 4645-4649 [PMID: 2162051] |
| Cultivation medium: | LB-Lennox + ampicillin (100 μg/ml) |
| Cultivation temperature: | 37°C |
| Biosafety level: | L1 |
| Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pDESTMB14-noGFP-unc54-pRPS-0-GAS-1-roGFP2-ORP-1 (LMBP 9708) is available at BCCM/GeneCorner. This plasmid was deposited by C. Vlaeminck and Prof. Dr B. Braeckman. |
Note: Up-to-date, validated data are enclosed with the biological material. Nevertheless, these data are a snapshot at a given moment; further updates are always possible.