Last data update: 24 January 2024 16:39 CET
Plasmid name: pSCGALST (LMBP 4266)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p4266.gb
(View with Genome Compiler) p4266.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Human ST6 beta-galactosamide alpha-2,6-sialyltranferase 1 (ST6GAL1) |
| Promoter: | Saccharomyces cerevisiae hybrid UDP-glucose 4-epimerase/iso-1-cytochrome C promoter (GAL10/CYC1) |
| Ribosome binding site: |
- |
| Terminator: | Saccharomyces cerevisiae 2 micron plasmid (2μ) FLP terminator Saccharomyces cerevisiae iso-1-cytochrome C terminator (CYC1) |
| Selection marker: | Ampicillin (amp) Saccharomyces cerevisiae URA3; auxotrophic LEU2 defective (leu2-d; auxotrophic) |
| Replicon: | Escherichia coli plasmid pMB1 origin Phage f1 origin Saccharomyces cerevisiae 2 micron plasmid origin (2μ); incl. region conferring stability (STB) |
| Host range: | Escherichia coli Saccharomyces cerevisiae; ura3(-) |
| Parental clone: | pSCGAL10-SN |
| Further information: | The plasmid was constructed by isolating a 4kb fragment, containing the human ST6GAL1 gene from a human hepatoma library, and cloning it into the NotI-SfiI opened pSCGAL10-SN vector. Expression of the human ST6GAL1 gene is controlled by the regulatable GAL10/CYC1 hybrid promoter from S. cerevisiae and the CYC1 and 2μ FLP terminators. The hybrid promoter sequence consists of the upstream activation sequence (UAS-G) of the GAL10 gene, the GAL10 promoter and the 5' untranslated leader sequence of CYC1 . The vector can be used in a c(+) yeast as a high-copy number extrachromosomal vector using the 2μ plasmid origin, but it can also be directed to integrate into the yeast chromosome at LEU2 (by linearising with AflII) or at URA3 (by linearising with ApaI). The vector contains three types of replication origin: 1) the origin of replication for E. coli; 2) the origin of replication of plasmid 2μ for extrachromosomal replication in S. cerevisiae; 3) the origin of replication of the single-stranded DNA phage f1, so that it can be rapidly switched between the plasmid and the phage mode (ssDNA) of replication. The latter requires infection with a helper phage, e.g. M13KO7. The defective LEU2 gene (leu2-d) is poorly expressed, because there is only 30 bp of upstream sequence left in this plasmid; the expression is probably controlled by a promoter within the 2μ sequences. Since the length of the 3' flanking region of the human ST6GAL1 gene could not be completely traced back, there is uncertainty as to the cutting frequency of the restriction enzymes, except for the sites that were analysed in the authenticity test. The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727335.1. The nucleotide sequence of the 4kb insert containing the human ST6GAL1 corresponds with the GenBank accession number NM_003032.2. Other name of the plasmid is pEMBLyex4-ST10. |
| EMBL Accession number: | NM_003032.2, view at GenBank LT727335.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 26/10/2012 |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: BamHI, BglI/NotI, EcoRV and PstI. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Prof. Dr R. Contreras(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: |
pSCGALST (LMBP 4266) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr R. Contreras . |
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.