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GeneCorner plasmid details

Last data update: 30 November 2020 04:30 CET

Plasmid name: pCAGGS-CrmA-T291R (LMBP 4577)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p4577.gb (View with Genome Compiler)
p4577.txt
p4577.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Cowpox virus cytokine response modifier A cDNA (crmA); 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: pCAGGScrmA
Further information: pCAGGS-CrmA-T291R is useful for highly efficient expression of mutated crmA under the control of the AG promoter and the human CMV-IE enhancer in various mammalian cells.
As compared to the wt crmA cDNA, the threonine (T) codon at position 291 was replaced by an arginine (R) codon, creating a unique NruI (Bsp68I) site and abolishing a PvuII site.
The crmA pointmutant can no longer inhibit caspase activity and consequently apoptosis.
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 LT727106.1.
The nucleotide sequence of the crmA cDNA corresponds with the EMBL Nucleotide Sequence Database accession number M14217.1.
EMBL Accession number: M14217.1, view at EMBL, GenBank, DDBJ
LT727106.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 26/02/2008
Sequence detail:
285                 290 291             295                 300
wt CrmA:     5' ... GAT.GTC.AAT.GAA.GAG.TAT.ACA.GAA.GCA.GCT.GCA.GCA.ACT.TGT.GCG.CTG ... 3'
                    Asp Val Asn Glu Glu Tyr Thr Glu Ala Ala Ala Ala Thr Cys Ala Leu
                                                     PvuII
                                                         PstI

                    285                 290 291             295                 300
crmA mutant: 5' ... GAT.GTC.AAT.GAA.GAG.TAT.CGC.GAA.GCG.GCT.GCA.GCA.ACT.TGT.GCG.CTG ... 3'
                    Asp Val Asn Glu Glu Tyr Arg Glu Ala Ala Ala Ala Thr Cys Ala Leu
                                            ^^^       ^  
                                          Bsp98I         PstI
                                          NruI

^: mutated nucleotide.
Punctuation indicates reading frame.
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: BamHI/PvuII, Bsp68I/NcoI, Bsu15I/HindIII, EcoRI, PstI/PvuI and XbaI.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by Prof. Dr P. Vandenabeele(1) (2). It was constructed by G. Van Loo(1) (2).
(1) Department for Molecular Biomedical Research, VIB, Ghent, Belgium
(2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium
Plasmid reference: -
Related plasmid reference: Tewari et al., Cell 81 (1995), 801-809 [PMID: 7774019]
Vercammen et al., J. Exp. Med. 187 (1998), 1477-1485 [PMID: 9565639]
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-CrmA-T291R (LMBP 4577) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr P. Vandenabeele .

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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