Last data update: 24 January 2024 16:39 CET
Plasmid name: pGMCSFlacZ (LMBP 2979)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p2979.gb
(View with Genome Compiler) p2979.txt p2979.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Escherichia coli lac Z gene (lacZ); modified N-terminal fragment |
| Promoter: | Mouse granulocyte-macrophage colony stimulating factor 2 promoter (Csf2; GMCSF; Gm-CSF) Escherichia coli lac operon promoter Phage T7 gene 10 promoter (T7g10) Phage T3 promoter |
| Ribosome binding site: |
- |
| Terminator: | Simian virus 40 polyadenylation signal (SV40 polyA) |
| Selection marker: | Ampicillin (amp) |
| Replicon: | Escherichia coli plasmid pMB1 origin Phage f1 origin |
| Host range: | Escherichia coli Mammalian cells |
| Parental clone: | pSVM-1; pUC18; pGEM-11Zf(+); pLGM-CSF12 |
| Further information: | The plasmid was constructed as follows: 1) the mouse Csf2 promoter was isolated from plasmid pLGM-CSF12 as an EcoRI-ScaI fragment and cloned into the EcoRI-HindII opened pUC18 vector; 2) from this intermediate construct, the mouse Csf2 promoter was isolated as an EcoRI-HindIII fragment and cloned into the EcoRI-HindIII opened pGEM-11Zf(+) vector; 3) from this second intermediate plasmid, the ScaI-HindIII fragment containing the mouse Csf2 promoter and part of the ampicillin resistance gene was isolated and ligated to the ScaI-HindIII pSVM-1 fragment containing the lacZ gene and the completing part of the ampicillin resistance gene. The lacZ gene lacks 25 N-terminal nucleotides and contains the N-terminal fragment (92 nucleotides) of the E. coli xanthine-guanine phosphoribosyltransferase (XGPRT) gene as well as a 86 bp fragment of the E. coli tryptophanyl tRNA synthetase gene. The cloned lacZ gene can be used as an efficient reporter gene for monitoring mouse Csf2 promoter activity. This plasmid was stably transfected into human TNF-receptor expressing PC60 cells and proved to be functional. When cloning a fragment downstream from the lac promoter it may be advisable to use lacI(q) strains in order to prevent fortuitous expression of a possibly noxious polypeptide. The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727417.1. The nucleotide sequence of the mouse Csf2 promoter and of the 5' flanking region correspond with the Genbank accession number NM_009969.4, except for some nucleotides of the Csf2 promoter: the TG in positions 156 and 157, the AG in positions 559 and 560, the T in position 638 and the G in position 922 of the promoter are missing and the A nucleotide in position 921 is replaced by a T. Other name of the plasmid is pSVmGMlacZ. |
| EMBL Accession number: | NM_009969.4, view at GenBank LT727417.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 12/04/2012 |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: BamHI, KpnI, PvuI and PvuII. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by B. Depuydt(1) (2), Dr P. Vandenabeele(1) (2) and Prof. Dr W. Fiers(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: |
pGMCSFlacZ (LMBP 2979) is available at BCCM/GeneCorner. This plasmid was deposited by B. Depuydt , Dr P. Vandenabeele and Prof. Dr W. Fiers . |
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.