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GeneCorner plasmid details

Last data update: 24 January 2024 16:39 CET

Plasmid name: pCD-F-bio-Ub-K27 (LMBP 6596)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner non-core plasmid
Depositor's sequence: p6596.gb
Sequence
analysis results
Genecorner:

-

Cloned DNA: Human ubiquitin B cDNA (UBB, Ub, HEL-S-50, GeneID 7314); mutated sequence
FLAG epitope tag; N-terminal
Biotinylation sequence (bio-tag); N-terminal
Promoter: Human cytomegalovirus immediate early promoter (CMV-IE) and enhancer
Phage T7 gene 10 promoter (T7g10)
Simian virus 40 early promoter (SV40 early)
Escherichia coli lac operon promoter
Ribosome
binding site:
-
Terminator: Bovine growth hormone polyadenylation signal (BGH polyA)
Simian virus 40 polyadenylation signal (SV40 polyA)
Selection marker: Ampicillin (amp)
Neomycin (neo; G418)
Replicon: Escherichia coli plasmid pMB1 origin
Phage f1 origin
Simian virus 40 bidirectional origin (SV40)
Host range: Escherichia coli
Mammalian cells; SV40 permissive cells
Parental clone: pCD-F-bio-MK-N149; pET-His-AU-Ub-K27
Further information: The plasmid was constructed by PCR amplifying the human UBB cds from pET-His-AU-Ub-K27 and ligating it as a BamHI/ApaI fragment into Ubiquitin was PCR amplified and ligated intopCD-F-bio-MK-N149, replacing the human MALT1 cds.
This plasmid is generated for high-affinity isolation of ubiquitinated proteins of a specific linkage. Biotinylation requires co-transfection of a mammalian expression vector with BirA. For even greater specificity, this plasmid can be combined with the corresponding His-tagged ubiquitin for 2D (first His-purification, then biotin precipitation) isolation of ubiquitinated proteins under denaturing conditions, for example for proteomic studies.
EMBL Accession number: -
Latest sequence update: 04/02/2010
Sequence detail:
Primers used to amplify Ubiquitin:

Forward: 5' TGAGGATCCT.ATG.CAG.ATC.TTC.GTG 
               BamHI   ***

Reverse: 5' TAAGGGCCC.CTA.CCC.ACC.TCT.GAG.AC
               ApaI   +++

***: start codon.
+++: termination codon.
Punctuation indicates reading frame.
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: -
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:

pCD-F-bio-Ub-K27 (LMBP 6596) is available at BCCM/GeneCorner. This plasmid was deposited by 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.

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