Last data update: 23 November 2023 13:39 CET
Plasmid name: pLenti6-tdTomato-V5 puro (LMBP 8400)
| New search | Print data sheet |
| Price category: | Cat. 2 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p8400.gb
(View with Genome Compiler) p8400.txt p8400.pdf |
| Sequence analysis results Genecorner: |
Sanger: 129499.fasta |
| Cloned DNA: |
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: | 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) 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) Chloramphenicol (cam) Puromycin (puro) |
| 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-V5 puro; ptdTomato |
| Further information: | The construction of this plasmid is described in De Groote et al. (2016). This is a lentiviral Gateway destination vector. 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. 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 pLenti6-tdTomato-V5 puro vector corresponds with the EMBL Nucleotide Sequence Database accession number LT009456.1. Other names of the plasmid are pLenti6(puro)-tdTomato-V5-DEST cl.11, pLenti6Fluo-puro and pLenti6-tdTomato-V5-puro. |
| EMBL Accession number: | LT009456.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 16/01/2019 |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: ApaI/SpeI, HindIII, PvuII and XhoI. This plasmid has also been fully sequenced but the NGS sequence data were unfortunately incomplete. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | The 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 |
| 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 (34 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: | - |
Refer in your Materials and Methods: |
pLenti6-tdTomato-V5 puro (LMBP 8400) is available at BCCM/GeneCorner. The 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.