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GeneCorner plasmid details

Last data update: 30 October 2020 04:11 CET

Plasmid name: pCAGGS-mCASP-12-C298A (LMBP 4573)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p4573.gb (View with Genome Compiler)
p4573.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Mouse cysteinyl aspartate specific proteinase 12 cDNA (caspase-12, CASP-12, Casp12); mutated sequence
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
Further information: The mouse cysteinyl aspartate specific proteinase 12 cDNA (caspase-12, mCASP-12) has been mutated by replacing a cysteine codon at amino acid position 298 by an alanine codon, resulting in the loss of the PstI site in the coding sequence.
pCAGGS-mCASP-12-C298A is useful for highly efficient expression of mutated mouse caspase-12 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 expressed protein is catalytic inactive.
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.
Name mentioned in Kalai et al. (2003) is pCAGGS-caspase-12 C298A.
EMBL Accession number: -
Latest sequence update: 29/08/2002
Sequence detail:
Nucleotide sequence of the mutated mCASP-12 cDNA:

                              298
wild type:     5' ... CAG.GCC.TGC.AGA.GGC.AGA ... 3'
                      Gln Ala Cys Arg Gly Arg
                            PstI
 
C298A mutant:  5' ... CAG.GCC.GCC.AGA.GGC.AGA ... 3'
                      Gln Ala Ala Arg Gly Arg
                              ^^
^: Mutated nucleotide.
Punctuation indicates reading frame.
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: HindIII, NcoI and PstI.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by M. Lamkanfi(1) (2), Dr M. Kalai(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: Kalai et al., J. Cell Biol. 162 (2003), 457-467 [PMID: 12885762]
Related plasmid reference: Van de Craen et al., FEBS Lett. 403 (1997), 61-69 [PMID: 9038361]
Restricted distribution: - BCCM MTA
Distributed as: H/P active culture or plasmid DNA
Host for distribution: Escherichia coli K12 MC1061F'
Host reference: -
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-mCASP-12-C298A (LMBP 4573) is available at BCCM/GeneCorner. This plasmid was deposited by M. Lamkanfi , Dr M. Kalai and Prof. Dr P. Vandenabeele and was published in Kalai et al., 2003.

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.

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