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pLVX-IRES-mCherry vector (V012723#)

Basic Vector Information

pLVX-IRES-mCherry is an HIV-1-based, lentiviral expression vector that allows the simultaneous expression of your protein of interest and mCherry in virtually any mammalian cell type, including primary cells. mCherry is a mutant fluorescent protein derived from the tetrameric Discosoma sp. red fluorescent protein, DsRed (1). The vector expresses the two proteins from a bicistronic mRNA transcript, allowing mCherry to be used as an indicator of transduction efficiency and a marker for selection by flow cytometry. Expression of the bicistronic transcript is driven by the constitutively active human cytomegalovirus immediate early promoter (PCMV IE) located just upstream of the MCS. An encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES), positioned between the MCS and mCherry, facilitates cap-independent translation of mCherry from an internal start site at the IRES/mCherry junction (1). pLVX-IRES-mCherry 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 RNA (2), leading to increased viral titers from packaging 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 (3). Finally, pLVX-IRES-mCherry 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 (4). The vector also contains a pUC origin of replication and an E. coli ampicillin resistance gene (Ampr) for propagation and selection in bacteria.

Basic Vector Information
Vector Name pLVX-IRES-mCherry Antibiotic Resistance Ampicillin
Length 8172 bp Type Viral Expression & Packaging Vectors
Source Clontech Selection Marker mCherry
Copy Number High copy number Promoter CMV
Cloning Method Enzyme Cut Expression Method Constiutive, Stable

pLVX-IRES-mCherry vector Vector Map

pLVX-IRES-mCherry vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       Exported                8172 bp ds-DNA     circular SYN 08-APR-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pLVX-IRES-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8172)
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8172
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     LTR             1..634
                     /note="3' LTR"
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /note="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /note="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    2028..2143
                     /note="cPPT/CTS"
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     enhancer        2201..2504
                     /note="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2505..2708
                     /note="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     misc_feature    2843..3415
                     /note="IRES"
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2866..3417
                     /note="IRES"
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     CDS             3417..4127
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /note="mCherry"
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     misc_feature    4141..4729
                     /note="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             complement(4612..4623)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /note="Factor Xa site"
                     /translation="IEGR"
     LTR             4936..5569
                     /note="3' LTR"
                     /note="3' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(5697..5713)
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5721..5737
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="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(5745..5775)
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    5790..5811
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding site"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(6099..6687)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6858..7718)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7719..7823)
                     /gene="bla"
                     /note="AmpR promoter"
     polyA_signal    7871..8005
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"

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