Last data update: 25 September 2020 04:24 CEST
Plasmid name: pLT-mCASP-1p30-m2 (LMBP 3949)
|Price category:||Cat. 1 (cf. price list)|
|Status:||GeneCorner non-core plasmid|
|Cloned DNA:||Mouse cysteinyl aspartate specific proteinase 1 cDNA (caspase-1, CASP-1, ICE, Casp1); p30 subunit C285S mutant|
|Promoter:||Phage λ major leftward promoter (λ PL)
Phage T7 gene 10 promoter (T7g10)
Phage T3 promoter
|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; pAS2p20p10ICEc/s|
|Further information:||The mouse caspase-1 coding sequence (p30ICE-C285S) contains point mutations that convert the Cys in the active site of the conserved QACRG sequence to Ser and create an additional NheI site.
The Cys conversion is described as C285S, according to the numbering of the codons in the human caspase-1 coding sequence, although the mutated mouse serine is located on position 284.
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 wild-type mouse mCASP-1cDNA corresponds with the Genbank accession number NM_009807.2.
Other name of the plasmid is pLT10T3p20p10Cys/Ser ICE.
|EMBL Accession number:||NM_009807.2, view at GenBank|
|Latest sequence update:||19/10/1999|
Sequence of the inactivated (mutated) proteolytic activity site: 280 284 ...GAC.AAG.CCC.AAG.GTG.ATC.ATT.ATT.CAA.GCT.AGC.CGT.GGA.GAG.AAA.CAA... Asp Lys Pro Lys Val Ile Ile Ile Gln Ala Ser Arg Gly Glu Lys Gln * * *-- Q A S R G NheI *: mutated nucleotide
|Authenticity test:||The plasmid still needs to be subjected to the authenticity test.|
|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
|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)|
|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.