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GeneCorner plasmid details

Last data update: 24 January 2024 16:39 CET

Plasmid name: pCAGGS-E-hA20-C624A-C627A-C775A-C779A (LMBP 6570)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner non-core plasmid
Depositor's sequence: not available
Sequence
analysis results
Genecorner:

-

Cloned DNA: Human TNF alpha induced protein 3 cDNA (TNFAIP3, A20, OTUD7C, GeneID 7128); mutated sequence
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-E-hA20-C624A-C627A; pCAGGS-E-hA20-C775A-C779A
Further information: pCAGGS-EhA20-C624A-C627A or pCAGGS-EhA20-C775A-C779A digested with SmaI results in a fragment of 6680 and 598. The small fragment of C624A-C627A was ligated to the large fragment of C775A-C779A to obtain pCAGGS-E-hA20-C624A-C627A-C775A-C779A.
This construct has 4 point mutations disrupting Zinc Finger 4 and 7. These Zinc fingers are needed for NF-kB inhibition.
The plasmid is useful for highly efficient expression of the fusion gene under the control of the AG promoter 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 lacIq strains in order to prevent fortuitous expression of a possibly noxious polypeptide.
EMBL Accession number: -
Latest sequence update: 22/01/2010
Authenticity test: The plasmid still needs to be subjected to the authenticity test.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by 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
Plasmid reference: Verhelst et al., EMBO J. 31 (2012), 3845-3855 [PMID: 23032186] [DOI: 10.1038/emboj.2012.240]
Related plasmid reference: Afonina et al., EMBO Rep. 17 (2016), 914-927 [PMID: 27113748] [DOI: 10.15252/embr.201642109]
Li et al., Biochim. Biophys. Acta 1793 (2009), 346-353 [PMID: 18952128]
Li et al., Biochim. Biophys. Acta 1783 (2008), 1140-1149 [PMID: 18329387]
Lin et al., J. Biol. Chem. 281 (2006), 2095-2130 [PMID: 16306043]
Natoli et al., J. Biol. Chem. 273 (1998), 31262-31272 [PMID: 9813034]
Wertz et al., Nature 430 (2004), 694-699 [PMID: 15258597]
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-E-hA20-C624A-C627A-C775A-C779A (LMBP 6570) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr R. Beyaert and was published in Verhelst et al., 2012.

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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