Last data update: 24 January 2024 16:39 CET
Plasmid name: pDAI-SceI-SacB (LMBP 10353)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner non-core plasmid |
| Depositor's sequence: | not available |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Saccharomyces cerevisiae endonuclease I-SceI cDNA (I-SceI) Bacillus subtilis levansucrase cDNA (sacB, GeneID 936413) |
| Promoter: | Escherichia coli dihydrofolate reductase promoter (P1) |
| Ribosome binding site: |
- |
| Terminator: | Escherichia coli rrnB operon T1 terminator Escherichia coli rrnB operon T2 terminator |
| Selection marker: | Tetracycline (tet) |
| Replicon: | Broad-host-range Gram-negative Bordetella bronchiseptica S87 plasmid pBBR1 replicon |
| Host range: | Escherichia coli Gram-negative bacterial strains |
| Parental clone: | pDAI-SceI |
| Further information: | The plasmid was constructed by cloning the SacB gene into the NheI site of pDAI-SceI. Plasmid pDAI-SceI-SacB expresses the I-SceI homing endonuclease under the control of the constitutive P1 promoter, derived from dhfrIIb (dehydrofolate reductase, responsible for trimethoprim resistance). The plasmid can be used in combination with pGPI-SceI (LMBP 10350) or pGPI-SceI-XCm (LMBP 10351), where I-SceI expression results in cleavage at the I-SceI recognition site of the genomically integrated pGPI-SceI or pGPI-SceI-XCm plasmid. This produces a double strand break in the cromosome, which stimulates the host recombination/repair machinery, resulting in gene deletion via allelic replacement. The plasmid also encodes the SacB gene to accelerate curing. |
| EMBL Accession number: | - |
| Latest sequence update: | 08/05/2017 |
| Authenticity test: | The plasmid still needs to be subjected to the authenticity test. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | The plasmid was deposited by Dr A. Sass(1). It was obtained from Prof. Dr M. Valvano(2). (1) Laboratory of Pharmaceutical Microbiology, Department of Pharmaceutical Analysis, Ghent University, Ghent, Belgium (2) Infectious Diseases Research Group, Department of Microbiology and Immunology, University of Western Ontario, London, Ontario, Canada |
| Plasmid reference: | Hamad et al., Appl. Environ. Microbiol. 76 (2010), 3170-3176 [PMID: 20348312] [DOI: 10.1128/AEM.03024-09] |
| Restricted distribution: | - BCCM MTA |
| Distributed as: | H/P 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 + tetracycline (10 μg/ml) |
| Cultivation temperature: | 37°C |
| Biosafety level: | L1 |
| Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pDAI-SceI-SacB (LMBP 10353) is available at BCCM/GeneCorner. The plasmid was deposited by Dr A. Sass and was published in Hamad et al., 2010. |
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.