Last data update: 24 January 2024 16:39 CET
Plasmid name: pEF6-hMALT1-E4A-P2A-Hyg (LMBP 9273)
| 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: |
Human MALT1 paracaspase cDNA (MALT1, MLT, PCASP1, paracaspase 1); mutated sequence Porcine teschovirus-1 2A peptide (P2A) |
| Promoter: | Human elongation factor 1α promoter (EF1α) Phage T7 gene 10 promoter (T7g10) Escherichia coli lac operon promoter |
| Ribosome binding site: |
- |
| Terminator: | Simian virus 40 polyadenylation signal (SV40 polyA) |
| Selection marker: | Hygromycin (hyg); in-frame translation Ampicillin (amp) |
| Replicon: | Escherichia coli plasmid pMB1 origin |
| Host range: | Escherichia coli Mammalian cells |
| Parental clone: | pEF6-P2A-Hyg; pCD-F-bio-MK-E4A |
| Further information: | The plasmid was constructed by PCR amplifying the mutant human MALT1 coding sequence from pCD-F-bio-MK-E4A and cloning it in the KpnI/SpeI opened pEF6-P2A-Hyg vector. The TRAF6 binding sites of human MALT1 were mutated. As compared to the wt human MALT1 ccoding sequence, the glutamic acid (E) codons at positions 313, 316, 653 and 806 were replaced by alanine (A) codons, creating additional SphI, AgeI and NruI sites and abolishing an Alw44I site. P2A is a self-cleaving peptide, fulfilling the same role as an IRES, but has a much shorter sequence. The purpose of this plasmid is to enforce expression of the human MALT1 mutant in selection-resistant cells by expressing the antibiotic resistance marker as an in-frame translation with your gene of interest, linked by the P2A autoprocessing peptide. The parental pEF6-P2A-Hyg vector lacks part of the pEF6 backbone, resulting in a deletion of among others the SV40 origin of replication. As a result, this plasmid is only suitable for stable expression after random genomic integration of the linearised plasmid. The nucleotide sequence of the wild type human MALT1 coding sequence corresponds with the EMBL Nucleotide Sequence Database accession number AF130356.2. |
| EMBL Accession number: | AF130356.2, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 12/04/2017 |
| 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 J. Staal(1)(2) and Prof. Dr R. Beyaert(1)(2). (1) VIB Center for Inflammation Research, Ghent, Belgium (2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium |
| Plasmid reference: | - |
| 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 + ampicillin (100 μg/ml) |
| Cultivation temperature: | 37°C |
| Biosafety level: | L1 |
| Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pEF6-hMALT1-E4A-P2A-Hyg (LMBP 9273) is available at BCCM/GeneCorner. The plasmid was deposited by Dr J. Staal and Prof. Dr R. Beyaert. |
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.