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LMBP plasmid details

Last data update: 04 December 2020 04:17 CET

Plasmid name: pLT-mCASP-1p30 (LMBP 4457)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p4457.gb (View with Genome Compiler)
p4457.txt
p4457.pdf
Sequence
analysis results
Genecorner:

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Cloned DNA: Mouse cysteinyl aspartate specific proteinase 1 cDNA (caspase-1, CASP-1, ICE, Casp1); p30 subunit
Promoter: Phage λ major leftward promoter (λ PL)
Phage T7 gene 10 promoter (T7g10)
Phage T3 promoter
Ribosome
binding site:
Ribosome binding site (RBS) of the phage T7 gene 10 (T7g10)
Terminator: Phage fd terminator
Phage T7 gene 10 terminator (T7g10)
Selection marker: Ampicillin (amp)
Replicon: Escherichia coli plasmid pMB1 origin
Phage f1 origin
Host range: Escherichia coli; use strains with a cI function, cIts for PL controlled expression
Parental clone: pLT10T3
Further information: The plasmid was created by cloning the mCASP-1 cDNA into the ApaI opened pLT10T3 plasmid.
This is a useful phasmid for mouse CASP-1p30 expression in E. coli under control of the strong and efficiently regulatable λ PL or T7 gene 10 promoter. Transcriptional read-through from these promoters is minimized by the presence of a duplicated T7 transcription terminator and a duplicated fd transcription terminator. Read-through transcription from other plasmid promoters is minimized by the clockwise orientation of the PL and T7 promoters relative to the anticlockwise orientation of the replication origin.
Furthermore, the phasmid is provided with an antisense phage T3 promoter. It also contains the replication origin of the single-stranded DNA phage f1, so that it can be rapidly switched between the plasmid and the phage mode (ssDNA) of replication.
For T7 driven expression use a strain containing a controllable T7 RNA polymerase gene, as well as a cI function: preferably a pT7POL plasmid (Mertens et al., Biotechnology 13 (1995), 175-179) or e.g. BL21(DE3)[pcI857]. Proceed as follows: first transform auxiliary plasmid, make competent cells again and then transform the expression plasmid.
The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727212.1.
The mCASP-1 cDNA was sequenced by BCCM/GeneCorner and corresponds with the EMBL Nucleotide Sequence Database accession number BC008152.1, except for some nucleotides in the 3' UTR.
Other name of the plasmid is pLT10T3p20p10.
EMBL Accession number: BC008152.1, view at EMBL, GenBank, DDBJ
LT727212.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 30/03/2009
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: AvaI, HincII, HindIII and NdeI/XhoI.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by Prof. Dr R. Beyaert(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(λ)
Host reference: Mertens et al., Gene 164 (1995), 9-15 [PMID: 7590329]
Related host reference: Casadaban et al., J. Mol. Biol. 138 (1980), 179-207 [PMID: 6997493]
Cultivation medium: LB-Lennox + ampicillin (100 μg/ml)
Cultivation temperature: 28°C
Biosafety level: L1
Cultivation remark: -
Other culture collection numbers: -

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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