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GeneCorner plasmid details

Last data update: 24 January 2024 16:39 CET

Plasmid name: pGAL1PNiST-1 (LMBP 4099)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p4099.gb (View with Genome Compiler)
p4099.txt
p4099.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Human interferon β gene (IFNB, IFNB1); stuffer fragment
Promoter: Candida albicans galactokinase 1 promoter (CaGAL1)
Escherichia coli lac operon promoter
Phage SP6 promoter
Phage T7 gene 10 promoter (T7g10)
Ribosome
binding site:
-
Terminator: Saccharomyces cerevisiae iso-1-cytochrome C terminator (CYC1)
Selection marker: Ampicillin (amp)
Candida albicans URA3; auxotrophic
Replicon: Escherichia coli plasmid pMB1 origin
Phage f1 origin
Host range: Escherichia coli
Candida albicans; ura3(-), integrative
Parental clone: pDBV50; pRM2; pPCK1NiSCT-1; pGEM-11Zf(+)
Further information: The plasmid was constructed as follows: 1) The EcoRI-SalI fragment from pDBV50, containing the C. albicans MAL2 promoter, was ligated into the EcoRI-SalI opened pGEM-11Zf(+) vector, resulting in the intermediate construct pGEMMAL2P-1. 2) pGEMMAL2P-1 was MscI opened and the C. albicans URA3 selection marker was cloned as an Eco47III-XmnI fragment from pRM2. 3) The resulting pGEMMAL2P-2 vector was NotI-HindIII opened in order to accept the EagI-HindIII fragment from pPCK1NiSCT-1, containing the NotI-stuffer-SfiI cassette, leading to pMAL2PNiST-1. 4) Finally, the SalI-EcI136II MAL2 promoter fragment in pMAL2PNiST-1 was exchanged by the XhoI-SmaI GAL1 promoter fragment from C. albicans, resulting in pGAL1PNiST-1.
Blasting the compiled nucleotide sequence of the plasmid against the Candida genome (http://www.candidagenome.org/), the (+-) 420 bp sequence upstream of the URA3 start codon corresponds to the Candida genomic DNA region between ORF 19.9283 (IRO1) and ORF 19.9284 (URA3), differing only in one nucleotide. Furthermore, Dr M. Logghe, who constructed the plasmid, confirmed that the URA marker is functional. Consequently, the plasmid carries, in all probability, the intact, native URA3 promoter region.
This integrative vector is useful for the construction of antisense cDNA expression libraries for C. albicans.
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 LT727304.1.
The nucleotide sequence of the C. albicans URA3 DNA corresponds with the EMBL Nucleotide Sequence Database accession number AF109400.1. This was confirmed by the sequence analysis results of BCCM/GeneCorner.
The nucleotide sequence of the C. albicans GAL1 promoter corresponds with the EMBL Nucleotide Sequence Database accession number Y13580.1.
The nucleotide sequence of the S. cerevisiae CYC1 terminator corresponds with the EMBL Nucleotide Sequence Database accession number M37696.1.
The nucleotide sequence of the human IFNβ cDNA corresponds with the EMBL Nucleotide Sequence Database accession number V00546.1.
EMBL Accession number: AF109400.1, view at EMBL, GenBank, DDBJ
V00546.1, view at EMBL, GenBank, DDBJ
Y13580.1, view at EMBL, GenBank, DDBJ
M37696.1, view at EMBL, GenBank, DDBJ
LT727304.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 16/03/2012
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: AgeI/NarI, EcoRI, HindIII, NotI and SfiI.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by Janssen Pharmaceutica N.V.(1). It was constructed by Dr M. Logghe(2) (3).
(1) Beerse, Belgium
(2) Department for Molecular Biomedical Research, VIB, Ghent, Belgium
(3) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium
Plasmid reference: De Backer et al., Yeast 15 (1999), 1609-1618 [PMID: 10572258]
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:

pGAL1PNiST-1 (LMBP 4099) is available at BCCM/GeneCorner. This plasmid was deposited by Janssen Pharmaceutica N.V. and was published in De Backer et al., 1999.

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.

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