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LMBP plasmid details

Last data update: 20 October 2018 04:23 CEST  


Name: pLenti6-V5-APEX blast   Add to cart
Accession number: LMBP 9184
Sequence file: Depositor's sequence: p9184.gb     View with Genome Compiler
Status: LMBP non-core plasmid
Cloned DNA: B gene of the control of cell death locus of the Escherichia coli F plasmid (ccdB, lethal gene)
Glycine max cytosolic ascorbate peroxidase 1 cDNA (APX1, APEX1, GeneID 553156); C-terminal
V5 epitope; C-terminal
Promoter: Rous sarcoma virus/human immunodeficiency virus hybrid long terminal repeat (RSV/HIV-1 5' LTR)
Human cytomegalovirus immediate early promoter (CMV-IE) and enhancer
Escherichia coli lac operon promoter
Simian virus 40 early promoter (SV40 early)
Phage T3 promoter
Phage T7 gene 10 promoter (T7g10)
Synthetic prokaryotic EM7 promoter
Escherichia coli lac operon promoter; mutant (lacUV5)
Ribosome
binding site:
-
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)
Blasticidin (bsd)
Chloramphenicol (cam)
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: pLenti6-Flag blast
Further information: The plasmid was constructed by cloning the V5 epitope and soybean APX1 coding sequence into the pLenti6-Flag blast vector, replacing the Flag epitope.
This is a lentiviral Gateway destination vector, 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.
Depending on the gene cloned in this vector, some leakage expression may be detected.
Because of the high risk for recombination, 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 pLenti6-Flag blast vector corresponds with the EMBL Nucleotide Sequence Database accession number LT009451.1.
Other name of the plasmid is pLenti6(bla) -V5 - APEX - dest.
EMBL Accession number: LT009451.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 07/08/2017
Authenticity test: The restriction enzyme pattern has still to be analysed.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: The plasmid was deposited by Prof. Dr M. Bertrand(1)(2).
(1) Center for Inflammation Research, VIB, Ghent, Belgium
(2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium
Plasmid reference: -
Related plasmid reference: De Groote et al., BioTechniques 60 (2016), 252-259 [PMID: 27177818] [DOI: 10.2144/000114417]
Restricted distribution: - BCCM MTA
Distributed as: plasmid DNA
Host for distribution: Escherichia coli K12xB DB3.1
Host reference: -
Related host reference: Bernard et al., J. Mol. Biol. 226 (1992), 735-745 [PMID: 1324324]
Cultivation medium: LB-Lennox + ampicillin (100 μg/ml) + chloramphenicol (25 μg/ml)*
Cultivation temperature: 28°C
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: -

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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