Last data update: 30 October 2020 04:11 CET
Plasmid name: pCAGGS-mCASP-1p10HA (LMBP 4347)
|Price category:||Cat. 1 (cf. price list)|
|Status:||GeneCorner core plasmid|
(View with Genome Compiler)
|Cloned DNA:||Mouse cysteinyl aspartate specific proteinase 1 cDNA (caspase-1, CASP-1, ICE, Casp1); p10 subunit
Influenza HA epitope encoding the haemagglutinin tagging peptide; C-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
|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; pCAGGS-mCASP-1|
|Further information:||The plasmid was constructed by inserting a PCR amplicon from pCAGGS-mCASP-1, containing the p10 subunit of the mouse cysteinyl aspartate specific proteinase 1 cDNA (mCASP-1) provided with the C-terminal HA epitope between the XhoI and MscI (BalI) sites of pCAGGS. The SmaI site at the 3' end of mCASP-1 is removed by a silent mutation.
pCAGGS-mCASP-1p10HA is useful for highly efficient expression of the p10 subunit of mouse caspase-1 under the control of the chicken β-actin/rabbit β-globin hybrid promoter (AG) and the human CMV-IE enhancer in various mammalian cells.
The AG promoter sequence consists of the chicken β-actin promoter, the first exon and part of the first intron (that seems to have a strong enhancer-like activity) linked to a rabbit β-globin fragment, consisting of a 3' part of the second intron (inclusive a branch point which is required for normal splicing reactions) and a 5' part of the third exon.
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 LT727195.1.
The nucleotide sequence of the mouse mCASP-1cDNA corresponds with the EMBL Nucleotide Sequence Database accession number BC008152.1.
Other name of the plasmid is pCAGGSp10casp-1#10.
|EMBL Accession number:||BC008152.1, view at EMBL, GenBank, DDBJ
LT727195.1, view at EMBL, GenBank, DDBJ
|Latest sequence update:||17/08/2006|
Nucleotide sequence at the borders of the insert: |--> p10 subunit |315 320 5'...TTCCTCGAG.ATG.GGC.ATT.AAG.AAG.GCC.CAT.ATA... XhoI 400 402| |<-- HA epitope -->| TTC.CCC.GGA.CAT.AGC.GGC.CGC.TAC.CCA.TAC.GAT.GTT.CCA.GAT.TAC.GCT.TAA.TGGCCAAT...3' * BalI *: Mutated nucleotide to remove the SmaI site (wild type = G). Punctuation indicates reading frame. Forward primer: 5' ...CACCGCTCGAGATGGGCATTAAGAAGGCCCATATAGAG...3' Reverse primer: 5'...CCATTAAGCGTAATCTGGAACATCGTATGGGTAGCGGCCGCTATGTCCGGGGAAGAGGTAGAAACGTTTTGTC...3'
|Authenticity test:||Restriction enzyme pattern analysed at GeneCorner: BsrDI, HindIII/XhoI and NotI.|
|History of deposit:||This plasmid was deposited by P. Schotte(1) (2) and Prof. Dr R. Beyaert (1) (2).
(1) Department for Molecular Biomedical Research, VIB, Ghent, Belgium
(2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium
|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)|
|Other culture collection numbers:||-|
Refer in your Materials and Methods:
|pCAGGS-mCASP-1p10HA (LMBP 4347) is available at BCCM/GeneCorner. This plasmid was deposited by P. Schotte 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.