Last data update: 24 January 2024 16:39 CET
Plasmid name: pCAGGS-EmCASP-8-C362A (LMBP 4942)
New search | Print data sheet |
Price category: | Cat. 1 (cf. price list) |
Status: | GeneCorner core plasmid |
GeneCorner sequence: |
p4942.gb
(View with Genome Compiler) p4942.txt p4942.pdf |
Sequence analysis results Genecorner: |
- |
Cloned DNA: |
Mouse cysteinyl aspartate specific proteinase 8 cDNA (caspase-8, CASP-8, Casp8, MCH5, MACH, FLICE); C362A mutant E-tag; N-terminal |
Promoter: | Chicken β-actin/rabbit β-globin hybrid promoter (AG) Human cytomegalovirus immediate early promoter (CMV-IE); enhancer only Escherichia coli lac operon promoter |
Ribosome binding site: |
- |
Terminator: | Rabbit β-globin polyadenylation signal (β-globin polyA) Simian virus 40 polyadenylation signal (SV40 polyA) |
Selection marker: | Ampicillin (amp) |
Replicon: | Escherichia coli plasmid pMB1 origin Simian virus 40 bidirectional origin (SV40) |
Host range: | Escherichia coli Mammalian cells; SV40 permissive cells |
Parental clone: | pCAGGS-HAmCASP-8-C362A; pCAGGS-EmCASP-1 |
Further information: | The plasmid was constructed by inserting the NotI/BamHI fragment of pCAGGS-HAmCASP-8-C362A between the NotI and BglII sites of pCAGGS-EmCASP-1. This plasmid carries a mouse caspase-8 cDNA mutant, with the cysteine codon in the catalytic site of the conserved QACQG sequence converted to an alanine codon (QAAQG), resulting in an additional MstI site. In Vanden Berghe et al. (2004), this C362A mutation is numbered C347A by mistake. pCAGGS-EmCASP-8-C362A is useful for highly efficient expression of mutated mouse caspase-8 under the control of the AG promoter and the human CMV-IE enhancer in various mammalian cells. The expressed protein is catalytically inactive. When cloning a fragment downstream from the lac promoter it may be advisable to use lacI(q) strains in order to prevent fortuitous expression of a possibly noxious polypeptide. The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727060.1. The nucleotide sequence of the wild type mouse CASP-8 cDNA corresponds with the EMBL Nucleotide Sequence Database accession number AJ007749.1. Other names of the plasmid are pCAGGS-E-proC8-C/A and pCAGGS-mCASP-8C/A. |
EMBL Accession number: | AJ007749.1, view at EMBL, GenBank, DDBJ LT727060.1, view at EMBL, GenBank, DDBJ |
Latest sequence update: | 27/10/2008 |
Sequence detail: | Nucleotide sequence at the 5' end, the mutated region and the 3' end of the mouse CASP-8 cDNA: ---------------------- E-tag ---------------------> 5' ... GAATTCCACC.ATG.GGT.GCG.CCG.GTG.CCG.TAT.CCG.GAC.CCG.CTG.GAA.CCG.CGT.GCG.GCC. EcoRI *** Gly Ala Pro Val Pro Tyr Pro Asp Pro Leu Glu Pro Arg Ala Ala NcoI NotI -------- mouse CASP-8 1 5 360 362 365 GCC.ATG.GAT.TTC.CAG.AGT ... ATC.TTT.TTC.ATT.CAG.GCT.GCG.CAA.GGA.AGT.AAC... Ala Met Asp Phe Gln Ser Ile Phe Phe Ile Gln Ala Ala Gln Gly Ser Asn *** ^^^ NcoI Q A A Q G MstI FspI mCASP-8 ->| 475 480 | ... AAG.CTC.TTC.TTC.CCT.CCC.TAA.GGATCT ... 3' Lys Leu Phe Phe Pro Pro +++ ***: start codon. +++: termination codon. ^ : mutated nucleotide. Punctuation indicates reading frame. |
Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: BglII, FspI and NotI. |
Class: | Recombinant plasmid |
Type: | Plasmid |
History of deposit: | This plasmid was deposited by Dr T. Vanden Berghe(1) (2) and Prof. Dr P. Vandenabeele(1) (2). (1) Department for Molecular Biomedical Research, VIB, Ghent, Belgium (2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium |
Plasmid reference: | Vanden Berghe et al., J. Biol. Chem. 279 (2004), 7925-7933 [PMID: 14668343] |
Related plasmid reference: | Maelfait et al., J. Exp. Med. 205 (2008), 1967-1973 [PMID: 18725521] [DOI: 10.1084/jem.20071632] |
Restricted distribution: | - BCCM MTA |
Distributed as: | H/P active culture or plasmid DNA |
Host for distribution: | Escherichia coli K12 MC1061 |
Host reference: | Casadaban et al., J. Mol. Biol. 138 (1980), 179-207 [PMID: 6997493] |
Related host reference: | Brigé et al., Biochem. J. 394 (2006), 335-344 [PMID: 16293111] |
Cultivation medium: | LB-Lennox + ampicillin (100 μg/ml) |
Cultivation temperature: | 37°C |
Biosafety level: | L1 |
Cultivation remark: | - |
Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pCAGGS-EmCASP-8-C362A (LMBP 4942) is available at BCCM/GeneCorner. This plasmid was deposited by Dr T. Vanden Berghe and Prof. Dr P. Vandenabeele and was published in Vanden Berghe et al., 2004. |
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.