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LMBP plasmid details

Last data update: 25 September 2020 04:24 CEST

Plasmid name: pLT-mCASP-1m1 (LMBP 3917)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner non-core plasmid
Depositor's sequence: p3917.gb
Sequence
analysis results
Genecorner:

-

Cloned DNA: Mouse cysteinyl aspartate specific proteinase 1 cDNA (caspase-1, CASP-1, ICE, Casp1); 2DE mutant
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 mouse caspase-1 coding sequence (p45ICE-2DE) contains point mutations, converting two Asp (D) amino acids of the prodomain to Glu (E) to destroy potential cleavage sites, and creating additional BalI (MscI) and KpnI restriction sites.
Transcriptional read-through from the 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.
The nucleotide sequence of the mouse mCASP-1cDNA corresponds with the EMBL Nucleotide Sequence Database accession number BC008152.1.
Other name of the plasmid is pLT10T3PROCASP-1Asp.
EMBL Accession number: BC008152.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 31/05/1999
Sequence detail:
Sequence of the mutated amino acids of the prodomain:

             100     102                             110
  ...TTT.GTT.GCT.ACA.GAA.GAA.TCT.AAA.GGT.GGC.CAT.CCT.TCA.TCC.TCA.GAA
     Phe Val Ala Thr Glu Glu Ser Lys Gly Gly His Pro Ser Ser Ser Glu
                     --*               *   *
                                       BalI (MscI)
 
                          prodomain --> | --> p30
                             120     122|
     ACA.AAG.GAA.GAA.CAG.AAC.AAA.GAA.GAG.GGT.ACC.TTT.CCA.GGA.CTG.ACT...
     Thr Lys Glu Glu Gln Asn Lys Glu Glu Gly Thr Phe Pro Gly Leu Thr
                                     --*   *   *
                                         KpnI
*: mutated nucleotide
Authenticity test: The plasmid still needs to be subjected to the authenticity test.
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: -
Related plasmid reference: Van Criekinge et al., J. Biol. Chem. 271 (1996), 27245-27248 [PMID: 8910297]
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
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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