Last data update: 24 January 2024 16:39 CET
Plasmid name: pCD-F-bio-Ub-K27 (LMBP 6596)
New search | Print data sheet |
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.