Last data update: 24 January 2024 16:39 CET
Plasmid name: pCD-F-bio-hUb (LMBP 6798)
New search | Print data sheet |
Price category: | Cat. 1 (cf. price list) |
Status: | GeneCorner non-core plasmid |
Depositor's sequence: | p6798.gb |
Sequence analysis results Genecorner: |
- |
Cloned DNA: |
Human ubiquitin B cDNA (UBB, Ub, HEL-S-50, GeneID 7314) 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; pET3E-His-AU1-hUb |
Further information: | The plasmid was constructed by PCR amplifying the human UBB coding sequence from pET3E-His-AU1-hUb and ligating it as a BamHI/ApaI fragment into pCD-F-bio-MK-N149, replacing the human MALT1 coding sequence. 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. The nucleotide sequence of the human UBB coding sequence corresponds with the EMBL Nucleotide Sequence Database accession number BT020104.1, except for some silent mutations. Other name of the plasmid is pCD-F-bio-Ub-wt. |
EMBL Accession number: | BT020104.1, view at EMBL, GenBank, DDBJ |
Latest sequence update: | 29/10/2010 |
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 Dr J. Staal(1) (2) and Prof. R. Beyaert(1) (2). (1) VIB-UGent Center for Inflammation 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 |
Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pCD-F-bio-hUb (LMBP 6798) is available at BCCM/GeneCorner. This plasmid was deposited by Dr J. Staal and Prof. 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.