Last data update: 22 May 2023 13:52 CEST
Plasmid name: pRMCE-DV2 (LMBP 8190)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner non-core plasmid |
| Depositor's sequence: | p8190.gb |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
B gene of the control of cell death locus of the Escherichia coli F plasmid (ccdB, lethal gene) |
| Promoter: | Mouse phosphoglycerate kinase 1 promoter (PGK1) Escherichia coli lac operon promoter Escherichia coli lac operon promoter; mutant (lacUV5) |
| Ribosome binding site: |
- |
| Terminator: | - |
| Selection marker: | Ampicillin (amp) Chloramphenicol (cam) |
| Replicon: | Escherichia coli plasmid pMB1 origin |
| Host range: | Escherichia coli; use a ccdB-resistant strain for propagation Mammalian cells |
| Parental clone: | pROSA26-DV2; pROSALUC |
| Further information: | The plasmid was constructed by replacing the 5' UTR and 3' UTR regions of mouse ROSA26 and the neomycin resistance gene in pROSA26-DV2 with the heterospecific FRT sites present in pROSALUC. A PGK-ATG sequence was included to ensure a proper trap-coupled RMCE reaction when targeted into mouse G4 ROSALUC ES cells. pRMCE-DV2 is a highly efficient Recombinase-Mediated Cassette Exchange (RMCE)-compatible, conditional Gateway destination vector. This multisite Gateway destination vector contains the ccdB gene inserted between the attenuator sites attR3 and attR4. The plasmid contains: - two heterospecific Saccharomyces cerevisiae 2μ FLP Recombinase Target sites (one wt and one mutant FRT site) - two adjacent copies of the chicken β-globin 5' DNase I hypersensitive site 4 (5'HS4) insulator core sequence. Insulators are DNA sequences which possess the ability to protect expressing genes from inappropriate signals emanating from their surrounding environment by acting as barriers that prevent the advance of nearby condensed chromatin that may otherwise silence expression. The resulting multisite Gateway expression vector, by co-transfection of a FlpE plasmid, can subsequently be targeted to the ROSA26 locus of G4 ROSALUC ES cells using a trap-coupled RMCE approach, hereby restoring neomycin resistance. The G4 ROSALUC ES cells (LMBP 10507CB) are available at BCCM/GeneCorner as well. Other name of the plasmid is pRMCE DV2. |
| EMBL Accession number: | - |
| Latest sequence update: | 01/12/2016 |
| Authenticity test: | The plasmid still needs to be subjected to the authenticity test. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Prof. Dr J. Haigh(1). (1) Australian Centre for Blood Diseases, Central Clinical School, Monash University, Melbourne, Australia |
| Plasmid reference: | PhD thesis Lieven Haenebalcke |
| Related plasmid reference: | Nyabi et al., Nucleic Acids Res. 37 (2009), e55 [PMID: 19279185] [DOI: 10.1093/nar/gkp112] |
| Restricted distribution: | - VIB/BCCM MTA - The depositor will be informed of the customer's identity upon release of a sample outside the depositor's department or outside the departments in which BCCM/GeneCorner is embedded, namely UGent-DBMB and VIB-IRC. |
| Distributed as: | 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: |
pRMCE-DV2 (LMBP 8190) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr J. Haigh and was published in Lieven Haenebalcke, . |
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.