Basic Vector Information
pLVX-tdTomato-N1 is an HIV-1-based, lentiviral expression vector that allows you to express your gene of interest fused to tdTomato, a member of the family of fruit fluorescent proteins (1) derived from the Discosoma sp. red fluorescent protein, DsRed (2). Genes cloned into the multiple cloning site (MCS), located upstream of the tdTomato coding sequence, are expressed as N-terminal tdTomato fusion proteins. Expression of the fusion protein is driven by the constitutively active human cytomegalovirus immediate early promoter (PCMV IE), located just upstream of the MCS. Lentiviral particles derived from the vector allow the expression of tdTomato fusion proteins in virtually any cell type, including primary cells. The unmodified vector expresses tdTomato, and may be used to produce marker virus to optimize infection protocols. pLVX-tdTomato-N1 contains all of the viral processing elements necessary for the production of replication-incompetent lentivirus, as well as elements to improve viral titer, transgene expression, and overall vector function. The woodchuck hepatitis virus posttranscriptional regulatory element (WPRE) promotes RNA processing events and enhances nuclear export of viral and transgene RNA (3), leading to increased viral titers from packaging cells, and enhanced expression of your gene of interest in target cells. In addition, the vector includes a Rev-response element (RRE), which further increases viral titers by enhancing the transport of unspliced viral RNA out of the nucleus (4). Finally, pLVX-tdTomato-N1 also contains a central polypurine tract/central termination sequence element (cPPT/CTS). During target cell infection, this element creates a central DNA flap that increases nuclear import of the viral genome, resulting in improved vector integration and more efficient transduction (5). In addition to lentiviral elements, pLVX-tdTomato-N1 contains a puromycin resistance gene (Puror) under the control of the murine phosphoglycerate kinase (PGK) promoter (PPGK) for the selection of stable transductants. The vector also contains a pUC origin of replication and an E. coli ampicillin resistance gene (Ampr) for propagation and selection in bacteria.
- Vector Name:
- pLVX-tdTomato-N1
- Antibiotic Resistance:
- Ampicillin
- Length:
- 9498 bp
- Type:
- Viral Expression & Packaging Vectors
- Replication origin:
- ori
- Source/Author:
- Clontech
- Selection Marker:
- Puromycin
- Copy Number:
- High copy number
- Promoter:
- mPGK
- Cloning Method:
- Enzyme Cut
- Fusion Tag:
- tdTomato
- Expression Method:
- Constiutive, Stable
pLVX-tdTomato-N1 vector Map
pLVX-tdTomato-N1 vector Sequence
LOCUS V012714 9498 bp DNA circular SYN 08-APR-2021
DEFINITION Exported.
ACCESSION V012714
VERSION V012714
KEYWORDS pLVX-tdTomato-N1
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 9498)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 9498)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..9498
/mol_type="other DNA"
/organism="synthetic DNA construct"
LTR 1..634
/label="3' LTR"
/note="3' long terminal repeat (LTR) from HIV-1"
misc_feature 681..806
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1303..1536
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 1721..1765
/label="gp41 peptide"
/note="antigenic peptide corresponding to amino acids 655
to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
al., 2013)"
CDS 1914..1955
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
misc_feature 2027..2144
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
enhancer 2201..2504
/label="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 2505..2708
/label="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
misc_feature 2793..2873
/label="MCS"
/note="multiple cloning site"
CDS 2875..4302
/label="tdTomato"
/note="tandem dimeric (pseudo-monomeric) derivative of
DsRed (Shaner et al., 2004)"
promoter 4333..4832
/label="PGK promoter"
/note="mouse phosphoglycerate kinase 1 promoter"
CDS 4853..5449
/label="PuroR"
/note="puromycin N-acetyltransferase"
misc_feature 5466..6054
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 6261..6894
/label="5' LTR"
/note="5' long terminal repeat (LTR) from HIV-1"
primer_bind complement(7023..7039)
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(7047..7063)
/label="lac operator"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
promoter complement(7071..7101)
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind complement(7116..7137)
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(7425..8013)
/direction=LEFT
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(8187..9044)
/label="AmpR"
/note="beta-lactamase"
promoter complement(9045..9149)
/label="AmpR promoter"
polyA_signal 9197..9331
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
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