pLVX-EF1α-IRES-mCherry

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  • Model: PVT2309
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pLVX-EF1α-IRES-mCherry

PVT2309    2ug

 

pLVX-EF1α-IRES-mCherry Informaiton

Promoter: EF-1 alpha promoter

Replicon: pUC ori

Terminator: SV40 poly (A) signal

Plasmid classification: virus series, lentivirus cloning vector

Prokaryotic resistance: ampicillin Amp

Selection marker: red fluorescence

Clone strain: Escherichia coli Stbl3

Culture conditions: 37 C, aerobic, LB

Expression host: lactation cells

Induction mode: no need to induce, transient expression.

5'sequencing primers: WPRE-R:CATAGCGTAAAAGGAGCAACA

Primers for 3'sequencing: primers designed based on sequences

Remarks: there is a difference from the sequence of Clontech.

 

 

pLVX-EF1α-IRES-mCherry Description

pLVX-EF1α-IRES-mCherry is a bigenic lentiviral expression vector that can be used to transduce mammalian cells to produce high titers of lentiviruses, including primary and stem cells, in almost any division or non-division. The vector contains an internal ribosome entry site (IRES) that allows an interesting gene and red fluorescent protein mCherry to be simultaneously transcribed from a single mRNA expression. Transcriptionally expressed human elongation factor-1 alpha (EF-1 alpha) promoters are driven and continue to be activated even when the genome of the host cell is stably integrated into the vector. Stable transcription allows the monitoring of various cellular processes, such as primary or stem cell differentiation, not to be silenced with CMV promoters. In addition, vectors allow efficient flow cytometry to detect the expressed proteins, mCherry, and benefits of stable or transient transfection of mammalian cells without time-consuming drug and cloning options.

 

pLVX-EF1α-IRES-mCherry Sequence

LOCUS       Exported                8935 bp ds-DNA     circular SYN 07-SEP-2016

DEFINITION  synthetic circular DNA.

ACCESSION   .

VERSION     .

KEYWORDS    pLVX-EF1-alpha-IRES-mCherry

SOURCE      synthetic DNA construct

  ORGANISM  synthetic DNA construct

REFERENCE   1  (bases 1 to 8935)

  AUTHORS   .

  TITLE     Direct Submission

  JOURNAL   Exported Wednesday, September 7, 2016 from SnapGene Viewer 3.1.4

FEATURES             Location/Qualifiers

     source          1..8935

                     /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)"

     promoter        2338..3519

                     /note="EF-1-alpha promoter"

                     /note="strong constitutive promoter for human elongation 

                     factor EF-1-alpha"

     intron          2568..3510

                     /note="EF-1-alpha intron A"

                     /note="intron upstream of the start codon of human 

                     EF-1-alpha"

     misc_feature    3575..4147

                     /note="IRES"

                     /note="internal ribosome entry site (IRES) of the 

                     encephalomyocarditis virus (EMCV)"

     CDS             4180..4890

                     /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    4904..5492

                     /note="WPRE"

                     /note="woodchuck hepatitis virus posttranscriptional 

                     regulatory element"

     CDS             complement(5375..5386)

                     /codon_start=1

                     /product="Factor Xa recognition and cleavage site"

                     /note="Factor Xa site"

                     /translation="IEGR"

     LTR             5699..6332

                     /note="3' LTR"

                     /note="3' long terminal repeat (LTR) from HIV-1"

     primer_bind     complement(6460..6476)

                     /note="M13 rev"

                     /note="common sequencing primer, one of multiple similar 

                     variants"

     protein_bind    6484..6500

                     /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(6508..6538)

                     /note="lac promoter"

                     /note="promoter for the E. coli lac operon"

     protein_bind    6553..6574

                     /bound_moiety="E. coli catabolite activator protein"

                     /note="CAP binding site"

                     /note="CAP binding activates transcription in the presence 

                     of cAMP."

     rep_origin      complement(6862..7450)

                     /direction=LEFT

                     /note="ori"

                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 

                     replication"

     CDS             complement(7621..8481)

                     /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(8482..8586)

                     /gene="bla"

                     /note="AmpR promoter"

     polyA_signal    8634..8768

                     /note="SV40 poly(A) signal"

                     /note="SV40 polyadenylation signal"

ORIGIN

        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca

       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac

      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca

      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg

      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag

      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg

      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat

      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga

      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct

      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc

      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag

      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg

      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga

      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg

      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg

      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct

      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat

     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca

     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc

     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag

     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc

     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg

     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc

     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc

     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct

     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa

     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca

     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt

     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt

     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta

     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt

     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct

     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga

     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccttt aaaagaaaag

     2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac

     2101 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg

     2161 acagcagaga tccagtttat cgatgagtaa ttcatacaaa aggactcgcc cctgccttgg

     2221 ggaatcccag ggaccgtcgt taaactccca ctaacgtaga acccagagat cgctgcgttc

     2281 ccgccccctc acccgcccgc tctcgtcatc actgaggtgg agaagagcat gcgtgaggct

     2341 ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag

     2401 gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg

     2461 tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag

     2521 tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt

     2581 gtggttcccg cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt

     2641 ccacgcccct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg

     2701 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg

     2761 cttgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct

     2821 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg

     2881 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc

     2941 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga

     3001 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct

     3061 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca

     3121 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg

     3181 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg

     3241 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat

     3301 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg

     3361 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa

     3421 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca

     3481 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gatctatttc cggtgaattc

     3541 ctcgagacta gttctagagc ggccgcggat cccgcccctc tccctccccc ccccctaacg

     3601 ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca

     3661 ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc ttcttgacga

     3721 gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg aatgtcgtga

     3781 aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg accctttgca

     3841 ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag

     3901 atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata gttgtggaaa

     3961 gagtcaaatg gctcacctca agcgtattca acaaggggct gaaggatgcc cagaaggtac

     4021 cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt gtttagtcga

     4081 ggttaaaaaa cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg

     4141 atgataagct tgccacaacc cacaaggaga cgaccttcca tggtgagcaa gggcgaggag

     4201 gataacatgg ccatcatcaa ggagttcatg cgcttcaagg tgcacatgga gggctccgtg

     4261 aacggccacg agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag

     4321 accgccaagc tgaaggtgac caagggtggc cccctgccct tcgcctggga catcctgtcc

     4381 cctcagttca tgtacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac

     4441 ttgaagctgt ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc

     4501 ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg

     4561 aagctgcgcg gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc

     4621 tgggaggcct cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgagatcaag

     4681 cagaggctga agctgaagga cggcggccac tacgacgctg aggtcaagac cacctacaag

     4741 gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca acatcaagtt ggacatcacc

     4801 tcccacaacg aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc

     4861 accggcggca tggacgagct gtacaagtga acgcgtctgg aacaatcaac ctctggatta

     4921 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg

     4981 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc

     5041 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca

     5101 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac

     5161 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact

     5221 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc

     5281 cgtggtgttg tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg

     5341 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc

     5401 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac

     5461 gagtcggatc tccctttggg ccgcctcccc gcctggaatt aattctgcag tcgagaccta

     5521 gaaaaacatg gagcaatcac aagtagcaat acagcagcta ccaatgctga ttgtgcctgg

     5581 ctagaagcac aagaggagga ggaggtgggt tttccagtca cacctcaggt acctttaaga

     5641 ccaatgactt acaaggcagc tgtagatctt agccactttt taaaagaaaa gaggggactg

     5701 gaagggctaa ttcactccca acgaagacaa gatatccttg atctgtggat ctaccacaca

     5761 caaggctact tccctgatta gcagaactac acaccagggc caggggtcag atatccactg

     5821 acctttggat ggtgctacaa gctagtacca gttgagccag ataaggtaga agaggccaat

     5881 aaaggagaga acaccagctt gttacaccct gtgagcctgc atgggatgga tgacccggag

     5941 agagaagtgt tagagtggag gtttgacagc cgcctagcat ttcatcacgt ggcccgagag

     6001 ctgcatccgg agtacttcaa gaactgctga tatcgagctt gctacaaggg actttccgct

     6061 ggggactttc cagggaggcg tggcctgggc gggactgggg agtggcgagc cctcagatcc

     6121 tgcatataag cagctgcttt ttgcctgtac tgggtctctc tggttagacc agatctgagc

     6181 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg

     6241 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag

     6301 acccttttag tcagtgtgga aaatctctag cagtagtagt tcatgtcatc ttattattca

     6361 gtatttataa cttgcaaaga aatgaatatc agagagtgag aggccttgac attgctagcg

     6421 tttaccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg

     6481 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc

     6541 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt

     6601 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg

     6661 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt

     6721 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc

     6781 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa

     6841 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa

     6901 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc

     6961 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc

     7021 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag

     7081 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga

     7141 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc

     7201 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac

     7261 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg

     7321 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca

     7381 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa

     7441 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa

     7501 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt

     7561 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag

     7621 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat

     7681 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc

     7741 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa

     7801 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca

     7861 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa

     7921 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt

     7981 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc

     8041 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact

     8101 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc

     8161 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg

     8221 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct

     8281 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc

     8341 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag

     8401 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac

     8461 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg

     8521 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt

     8581 tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atcaacttgt

     8641 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag

     8701 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg

     8761 tctggatcaa ctggataact caagctaacc aaaatcatcc caaacttccc accccatacc

     8821 ctattaccac tgccaattac ctgtggtttc atttactcta aacctgtgat tcctctgaat

     8881 tattttcatt ttaaagaaat tgtatttgtt aaatatgtac tacaaactta gtagt

//

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