Last data update: 21 October 2021 10:13 CEST
Plasmid name: pDG1i-tdTomato-V5 blast (LMBP 7942)
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|Price category:||Cat. 3 (cf. price list)|
|Status:||GeneCorner core plasmid|
(View with Genome Compiler)
B gene of the control of cell death locus of the Escherichia coli F plasmid (ccdB, lethal gene)|
Discosoma sp. red fluorescent protein DNA (DsRed1); mutated variant (tandem dimer Tomato, tdTomato)
V5 epitope; C-terminal
|Promoter:||Human cytomegalovirus immediate early promoter (CMV-IE); minimal promoter with tet responsive element (CMVTRE)
Rous sarcoma virus/human immunodeficiency virus hybrid long terminal repeat (RSV/HIV-1 5' LTR)
Simian virus 40 early promoter (SV40 early)
Escherichia coli lac operon promoter
Phage T3 promoter
Phage T7 gene 10 promoter (T7g10)
Synthetic prokaryotic EM7 promoter
Escherichia coli lac operon promoter; mutant (lacUV5)
|Terminator:||Rous sarcoma virus/human immunodeficiency virus hybrid long terminal repeat (RSV/HIV-1 3' LTR); modified HIV-1 3' LTR with deleted U3 region (ΔU3/3' LTR)
Simian virus 40 polyadenylation signal (SV40 polyA)
|Selection marker:||Ampicillin (amp)
|Replicon:||Escherichia coli plasmid pMB1 origin
Phage f1 origin
Simian virus 40 bidirectional origin (SV40)
|Host range:||Escherichia coli; use a ccdB-resistant strain for propagation
Mammalian cells; SV40 permissive cells
|Parental clone:||ptdTomato; pDG1i-V5 blast|
|Further information:||The construction of this plasmid is described in De Groote et al. (2016).
This is a lentiviral Gateway destination vector, with tet-inducible minimal human CMV-IE promotor.
The plasmid is suitable for microRNA expression.
The U3 region of the HIV-1 3' LTR is deleted (ΔU3 or dU3) and facilitates self-inactivation of the HIV-1 5' LTR after transduction to enhance the biosafety of the vector. The element also contains a polyadenylation signal for transcription termination and polyadenylation of mRNA in transduced cells.
The vector contains the psi packaging signal, allowing viral packaging, and the rev response element (RRE) from HIV-1, permitting Rev-dependent nuclear export of unspliced viral mRNA.
The red fluorescent protein tdTomato was derived from a monomeric mutant of DsRed by several rounds of directed mutagenesis and is detectable by DsRed antibodies.
Depending on the gene cloned in this vector, some leakage expression may be detected.
Because of the high risk for recombination, this plasmid is only available under the format of isolated plasmid DNA and plasmid integrity should be checked via PvuII restriction digest after each subcultivation.
This Gateway destination vector is compatible with the human orfeome library which is also available at BCCM/GeneCorner.
This vector can not be used to express short hairpin RNA (shRNA), which requires an RNA-polymerase III promotor. However, this vector can be used for knock-down experiments if the hairpin is expressed in a micro-RNA context (eg. Block-iT PolII miR RNAi kit).
The nucleotide sequence of the pDG1i-tdTomato-V5 blast vector corresponds with the EMBL Nucleotide Sequence Database accession number LT009440.1 except for the adjustments due to the implementation of the next-generation sequencing results.
Other names of the plasmid are pDGlenti(Blast)tdTomatoV5-Dest, pDG1iFluo-blast and pDG1i-tdTomato-V5-blast.
|EMBL Accession number:||LT009440.1, view at EMBL, GenBank, DDBJ|
|Latest sequence update:||17/01/2019|
|Authenticity test:||Restriction enzyme pattern analysed at GeneCorner: ApaI/SpeI, HindIII, PvuII and XhoI.
This plasmid has also been fully sequenced.
|History of deposit:||This plasmid was deposited by Dr P. De Groote(1) (2) and Prof. Dr W. Declercq(1) (2).
(1) Inflammation Research Center, VIB, Ghent, Belgium
(2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium
|Plasmid reference:||De Groote et al., BioTechniques 60 (2016), 252-259 [PMID: 27177818] [DOI: 10.2144/000114417]
|Related plasmid reference:||PhD thesis Philippe De Groote (2012)
|Restricted distribution:||- BCCM MTA|
|Distributed as:||plasmid DNA|
|Host for distribution:||Escherichia coli K12xB DB3.1|
|Related host reference:||Bernard et al., J. Mol. Biol. 226 (1992), 735-745 [PMID: 1324324]
|Cultivation medium:||LB-Lennox + ampicillin (100 μg/ml) + chloramphenicol (34 μg/ml)*|
|Biosafety level:||L1 in E. coli; L2 in mammalian cells|
|Cultivation remark:||*: selection of transformants on chloramphenicol; subsequent cultivation of a single colony in liquid medium with ampicillin|
|Other culture collection numbers:||-|
Refer in your Materials and Methods:
|pDG1i-tdTomato-V5 blast (LMBP 7942) is available at BCCM/GeneCorner. This plasmid was deposited by Dr P. De Groote and Prof. Dr W. Declercq and was published in De Groote et al., 2016.|
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.