LOCUS       sequence-259750-        6385 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NRF2 reporter plasmid with GFP under control of 8 ARE elements and 
            constitutively transcribed BFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6385)
  AUTHORS   Wyler E, Franke V, Menegatti J, Kocks C, Boltengagen A, Praktiknjo 
            S, Walch-Ruckheim B, Bosse J, Rajewsky N, Grasser F, Akalin A, 
            Landthaler M
  TITLE     Single-cell RNA-sequencing of herpes simplex virus 1-infected cells 
            connects NRF2 activation to an antiviral program.
  JOURNAL   Nat Commun. 2019 Oct 25;10(1):4878. doi: 10.1038/s41467-019-12894-z.
  PUBMED    31653857
REFERENCE   2  (bases 1 to 6385)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2019-10-25"; volume: "10(1):4878. doi"; pages: " 
            10.1038/s41467-019-12894-z"
COMMENT     Sequence Label: pREP-8xARE-GFP-SV40-BFP
FEATURES             Location/Qualifiers
     source          1..6385
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        12..328
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      179..314
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             425..1024
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(425..444)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     921..941
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1254..1335
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1590..1947
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1590..1781
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1613..1633)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1798..1933
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1969..2670
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     polyA_signal    2694..2815
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2705..2724)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             join(2976..3044,3045..3836)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3194..3213)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4007..4595
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4496..4515
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4749..4766
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4883..4904
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4919..4924,4925..4942,4943..4949)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4957..4973
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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)."
     primer_bind     4962..4984
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4981..4997
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4981..4997
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        5017..5035
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5017..5034
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     5047..5066
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5047..5065
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    5126..5174
                     /note="synthetic polyadenylation signal"
     misc_feature    5188..5279
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     5228..5247
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        5408..5439
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      5472..5481
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(5478..5480,5481..5483,5484..6197)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(5478..5480,5481..5483,5484..6194)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5523..5544)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5784..5803)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6131..6152
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(6216..6235)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6335..6357
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaagat tggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
       61 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
      121 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
      181 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
      241 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
      301 cttttttgga ggcctaggct tttgcaaaaa gctagcttgc atgcctgcag gtcggccgcc
      361 acgaccggtg ccgccaccat cccctgaccc acgcccctga cccctcacaa ggagacgacc
      421 ttccatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tcccccgggc
      481 cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgaccc
      541 ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct
      601 cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc
      661 ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag
      721 cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa
      781 ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct
      841 gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt
      901 cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac
      961 cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc
     1021 ctgacgcccg ccccacgacc cgcagcgccc gaccgaaagg agcgcacgac cccatggctc
     1081 cgaccgaagc cacccggggc ggccccgccg accccgcacc cgcccccgag gcccaccgac
     1141 tctagaggat cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc
     1201 tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt
     1261 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     1321 catttttttc actgcaatct cgtgatacgc ctatttttat aggttaatgt catgataata
     1381 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1441 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1501 cttcaataat atatctcgtc atgtgggccc atctgcgcag caccatggcc tgaaataacc
     1561 tctgaaagag gaacttggtt aggtaccttc tgaggcggaa agaaccagct gtggaatgtg
     1621 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     1681 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     1741 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     1801 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     1861 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
     1921 ttttttggag gcctaggctt ttgcaaaaag cttgattctt ctgacacgat gagcgagctg
     1981 attaaggaga acatgcacat gaagctgtac atggagggca ccgtggacaa ccatcacttc
     2041 aagtgcacat ccgagggcga aggcaagccc tacgagggca cccagaccat gagaatcaag
     2101 gtggtcgagg gcggccctct ccccttcgcc ttcgacatcc tggctactag cttcctctac
     2161 ggcagcaaga ccttcatcaa ccacacccag ggcatccccg acttcttcaa gcagtccttc
     2221 cctgagggct tcacatggga gagagtcacc acatacgaag acgggggcgt gctgaccgct
     2281 acccaggaca ccagcctcca ggacggctgc ctcatctaca acgtcaagat cagaggggtg
     2341 aacttcacat ccaacggccc tgtgatgcag aagaaaacac tcggctggga ggccttcacc
     2401 gagacgctgt accccgctga cggcggcctg gaaggcagaa acgacatggc cctgaagctc
     2461 gtgggcggga gccatctgat cgcaaacatc aagaccacat atagatccaa gaaacccgct
     2521 aagaacctca agatgcctgg cgtctactat gtggactaca gactggaaag aatcaaggag
     2581 gccaacaacg agacctacgt cgagcagcac gaggtggcag tggccagata ctgcgacctc
     2641 cctagcaaac tggggcacaa gcttaattaa gcggccgctt cgagcagaca tgataagata
     2701 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     2761 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     2821 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag
     2881 caagtaaaac ctctacaaat gtggtaaaat cgataaggat cgatccaggt ggcactttgg
     2941 gcccatctgc tgtcatatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     3001 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     3061 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     3121 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     3181 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     3241 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     3301 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     3361 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     3421 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     3481 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     3541 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     3601 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     3661 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     3721 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     3781 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     3841 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     3901 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     3961 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     4021 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttttt
     4081 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     4141 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     4201 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     4261 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     4321 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     4381 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     4441 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     4501 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     4561 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     4621 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     4681 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     4741 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     4801 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     4861 cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt
     4921 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
     4981 caggaaacag ctatgacatg attacgaatt gcaacgattt aggtgacact atagaagaga
     5041 aggaattaat acgactcact atagggagag agagagaatt accctcacta aagggaggag
     5101 aagcatgaat tcggcctaac tggccaataa aatatcttta ttttcattac atctgtgtgt
     5161 tggttttttg tgtgaatcga tagtactaac atacgctctc catcaaaaca aaacgaaaca
     5221 aaacaaacta gcaaaatagg ctgtccccag tgcaagtgca ggtgccagaa catttctctg
     5281 gcctaactgg ccgtgacact gcagcagtga ctacgcatgc gtgacgatgc agacgtgaca
     5341 ctgcactggt gacaaagcac ccgtgactag gcaagcgtga cgtagcatcg gtgacacagc
     5401 aagacactag agggtatata atggaagctc gacttccagc ttggcaatcc ggtactgttg
     5461 gtaaactcga ggccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca
     5521 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     5581 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     5641 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     5701 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     5761 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     5821 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     5881 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     5941 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     6001 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     6061 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     6121 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     6181 acgagctgta caagtaacat atggtgcact ctcagtacaa tctgctctga tgccgcatag
     6241 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     6301 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     6361 tcaccgtcat caccgaaacg cgcga
//
