Last data update: 24 January 2024 16:39 CET
Plasmid name: pGal4-p65 (LMBP 3438)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p3438.gb
(View with Genome Compiler) p3438.txt p3438.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Saccharomyces cerevisiae GAL4 DNA-binding domain (GALbd) Human nuclear factor κB cDNA (NF-κB); p65 (RELA) subunit |
| Promoter: | Rous sarcoma virus long terminal repeat (RSV-LTR) Simian virus 40 early promoter (SV40 early) |
| Ribosome binding site: |
- |
| Terminator: | 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: | pABgal; p65/3 |
| Further information: | This plasmid was constructed as follows: 1) p65/3, a Bluescript vector containing full-length hNF-κB p65 cDNA, was cut with SmaI; 2) the 1.6 kb SmaI fragment, containing the first 519 codons of the hNF-κB p65 coding sequence, was inserted in the PvuII site of the polylinker of pABgal; 3) the intermediate construct was then opened with BstEII-HindIII, followed by the insertion of the missing hNF-κB p65 cDNA fragment from p65/3. The plasmid contains the S. cerevisiae GALbd domain, composed of the first 147 codons from GAL4, fused to the full-length hNF-κB p65 coding sequence. pGal4-p65 is used as an activator plasmid in combination with a Gal4-dependent reporter plasmid (e.g. pGal4-CAT). The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727488.1. The nucleotide sequence was compiled in accordance with the description in Schmitz et al. (1991). The fusion region between GALbd and the hNF-κB p65 subunit was sequenced (F. Molemans, Dept of Biomedical Molecular Biology, Ghent University, Belgium). The nucleotide sequence of the hNF-κB p65 coding sequence corresponds with the EMBL Nucleotide Sequence Database accession number M62399.1. The nucleotide sequence of the 3' UTR of the hNF-κB p65 subunit was retrieved from the pRc/CMV-p65 sequence file provided by S. Plaisance (Dept of Biomedical Molecular Biology, Ghent University, Belgium). The nucleotide sequence upstream of GALbd (from nucleotide position 3841 up to and including 4920) was analysed by BCCM/GeneCorner. Name mentioned in Schmitz et al. (1991) is GAL4-p65 1-550. Other names of the plasmid are Gal4-p65-long and pAB65. |
| EMBL Accession number: | M62399.1, view at EMBL, GenBank, DDBJ LT727488.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 22/09/2003 |
| Sequence detail: | Nucleotide sequence at the start of the S. cerevisiae GAL4 DNA-binding domain:
---------- GALbd ------------->
2
5' ... ACTG.ATG.GAC.TCC.CAG.CAG.CCA.GAT.CTG.AAG.CTA.CTG.TCT.TCT.ATC.GAA ... 3'
Met Asp Ser Gln Gln Pro Asp Leu Lys Leu Leu Ser Ser Ile Glu
*** BglII
Nucleotide sequence at the GALbd-p65 fusion:
----------------- GALbd -------------->
147 ---------------- linker
5' ... AAA.GGT.CAA.AGA.CAG.TTG.ACT.GTA.TCG.CCG.GAA.TTC.CCG.GGT.GTC
Lys Gly Gln Arg Gln Leu Thr Val Ser Pro Glu Phe Pro Gly Val
HindII EcoRI AvaI HindII
SmaI SalI
--------------- p65 ------------------>
linker ---------------> 1
GAC.CAG.GGG.ACC.CCG.GCC.ATG.GAC.GAA.CTG.TTC.CCC.CTC.ATC.TTC ... 3'
Asp Gln Gly Thr Pro Ala Met Asp Glu Leu Phe Pro Leu Ile Phe
***
NcoI
StyI
*** : start codon.
Punctuation indicates reading frame. |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: BamHI, BamHI/XmnI, BstEII/HindIII, EcoRI, KpnI/SfiI, KpnI/SmaI, PvuI/XhoI and XhoI. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Dr M.L. Schmitz(1). (1) Albert-Ludwigs-Universit“t Freiburg, Germany |
| Plasmid reference: | Schmitz et al., EMBO J. 10 (1991), 3805-3817 [PMID: 1935902] |
| Related plasmid reference: | Campa et al., Oncotarget 5 (2014), 8173-8187 [PMID: 25327559] [DOI: 10.18632/oncotarget.2303] Chiloeches et al., Mol. Endocrinol. 22 (2008), 2466-2480 [PMID: 18755855] [DOI: 10.1210/me.2008-0107] Larsen et al., Diabetologia 48 (2005), 2582-2590 [PMID: 16283237] [DOI: 10.1007/s00125-005-0039-9] Lasa et al., Mol. Endocrinol. 24 (2010), 412-422 [PMID: 20032197] [DOI: 10.1210/me.2009-0298] Vanden Berghe et al., J. Biol. Chem. 273 (1998), 3285-3290 [PMID: 9452444] |
| Restricted distribution: | - BCCM MTA |
| Distributed as: | H/P active culture or plasmid DNA |
| Host for distribution: | Escherichia coli K12 DH5α |
| Host reference: | Focus 8 (1986), 9 |
| Related host reference: | Woodcock et al., Nucleic Acids Res. 17 (1989), 3469-3478 [PMID: 2657660] Rodriguez-Quinones et al., Focus 15 (1993), 110-112 Hanahan, J. Mol. Biol. 166 (1983), 557-580 [PMID: 6345791] [DOI: 10.1016/s0022-2836(83)80284-8] Hanahan, in 'DNA Cloning: A Practical Approach Volume I', IRL Press, Oxford (1985), 109-135 [ISSN/ISBN: 0947946187] Grant et al., Proc. Natl. Acad. Sci. U.S.A. 87 (1990), 4645-4649 [PMID: 2162051] |
| 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: |
pGal4-p65 (LMBP 3438) is available at BCCM/GeneCorner. This plasmid was deposited by Dr M.L. Schmitz and was published in Schmitz et al., 1991. |
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.