Last data update: 24 January 2024 16:39 CET
Plasmid name: pHis-p65 (LMBP 4650)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p4650.gb
(View with Genome Compiler) p4650.txt p4650.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Human nuclear factor κB cDNA (NF-κB); p65 (RELA) subunit Histidine tag (His-tag); N-terminal Xpress epitope; N-terminal Enterokinase cleavage site (EK); 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) |
| 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: | pHis-p65f2-286; pGal4-p65f286-550 |
| Further information: | The plasmid was constructed by inserting the EcoRI fragment of pGal4-p65f286-550, coding for the amino acids 285 up to and including 551 of the human NF-κB p65 subunit, into the EcoRI opened pHis-p65f2-286 vector. As a result, the full-length p65 subunit was fused in phase to the N-terminal histidine tag and Xpress‚ epitope. The plasmid is designed for high-level expression of the tagged hNF-κB p65 subunit in mammalian cells, under control of the strong hCMV promoter. The parental clone of this vector has been shown to provide resistance to kanamycin, although growth was reduced compared to growth on medium containing ampicillin. The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727144.1. The nucleotide sequence of the human NF-κB p65 coding sequence corresponds with the EMBL Nucleotide Sequence Database accession number M62399.1. The nucleotide sequence of the 3' UTR of the human NF-κB p65 cDNA was retrieved from the pRc/CMV-p65 sequence file provided by S. Plaisance (Dept of Biomedical Molecular Biology, Ghent University, Belgium). |
| EMBL Accession number: | M62399.1, view at EMBL, GenBank, DDBJ LT727144.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 30/10/2003 |
| Sequence detail: | Nucleotide sequence at the start of the tagged hNF-kB p65 subunit:
------- His-tag ------>
5' ... TTAAGCTTACC.ATG.GGG.GGT.TCT.CAT.CAT.CAT.CAT.CAT.CAT.GGT.ATG.GCT.AGC
HindIII Met Gly Gly Ser His His His His His His Gly Met Ala Ser
*** NheI
NcoI
StyI
------- Xpress epitope ------->
- EK recogn.site ->
ATG.ACT.GGT.GGA.CAG.CAA.ATG.GGT.CGG.GAT.CTG.TAC.GAC.GAT.GAC.GAT.AAG
Met Thr Gly Gly Gln Gln Met Gly Arg Asp Leu Tyr Asp Asp Asp Asp Lys
KpnI
--------------------------- p65 --------------------------->
2
GTA.CCG.GAC.GAA.CTG.TTC.CCC.CTC.ATC.TTC.CCG.GCA.GAG.CCA.GCC.CAG.GCC ... 3'
Val Pro Asp Glu Leu Phe Pro Leu Ile Phe Pro Ala Glu Pro Ala Gln Ala
Nucleotide sequence at the end of the hNF-kB p65 subunit:
--------------------------- p65 --------------------------->
551
5' ... GCG.GAC.ATG.GAC.TTC.TCA.GCC.CTG.CTG.AGT.CAG.ATC.AGC.TCC.TAA.GGGGGTG ... 3'
Ala Asp Met Asp Phe Ser Ala Leu Leu Ser Gln Ile Ser Ser +++
***: start codon.
+++: termination codon.
Punctuation indicates reading frame. |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: BanII, EcoRI, EcoRV/KpnI, HindII, PstI, StuI and XhoI. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Dr L. Vermeulen(1) and Prof. Dr G. Haegeman(1). (1) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium |
| Plasmid reference: | Reber et al., PLoS One. 4 (2009), e4393 [PMID: 19197368] [DOI: 10.1371/journal.pone.0004393] |
| 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: |
pHis-p65 (LMBP 4650) is available at BCCM/GeneCorner. This plasmid was deposited by Dr L. Vermeulen and Prof. Dr G. Haegeman and was published in Reber et al., 2009. |
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.