LOCUS       sequence-479907-        7254 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express NRF2-SmBiT_MAFG-LgBiT Fusion Protein in Mammalian Cells 
            under a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7254)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7254)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: NRF2-SmBiT_MAFG-LgBiT BiBiT Vector
FEATURES             Location/Qualifiers
     source          1..7254
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    24..105
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        complement(1337..1355)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1338..1355)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        1475..1778
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1779..1982
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1933..1953
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          2118..2250
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2233..2252
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     2294..2313
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2294..2312
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    4317..4438
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4328..4347)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(4553..4586)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        4596..4643
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4675..5469
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4729..4748)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5339..5358
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(5627..6215)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5707..5726)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    6341..6389
                     /note="synthetic polyadenylation signal"
     CDS             complement(6413..6811)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      6808..6817
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(6842..7199)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6856..6991
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        7008..7199
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     7156..7176
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 aagacgcgtt aattaagacg tcggcagtga aaaaaatgct ttatttgtga aatttgtgat
       61 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
      121 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
      181 ctctacaaat gtggtatggc tgattatgat ccgcggccgc aatacgtcga cgttatcagc
      241 tgacttcgta cgagagccta ggtgtttcta gagtcgcggc cttagctgtt gatggttact
      301 cggaacagca tggagccgtc gggggtgatc aggcgctcgt cgataatttt gttgccgttc
      361 cacagggtcc ctgttacagt gatctttttg ccgtcgaaca cggcgatgcc ttcatacggc
      421 cgtccgaaat agttcagcat gttcggcgta accccgtcga ttaccagtgt gccatagggc
      481 aggatcacct taaagtgatg atcatccaca gggtacacca ccttaaacac ctcttcgatc
      541 tgggccattt ggtcggcgct cagaccttca tacgggatga tgacatggat gtcgatcttc
      601 agggcatttt caccgctccg gacaatcctt tggatcggag ttacggacac ggcgagattc
      661 tgcagcaaac tggacacacc tccctgttca aggacttggt ccaggttgta ggcggctgtc
      721 tgttcccagt ccccaacgaa atcttcgagt gtgaagacac cgctcgagcc tccacctccg
      781 ctcccgccac cacctgaact gccttgagaa accgatcggg catccgtctt ggactttact
      841 attgtgatga cgctggtggc ggccaccttg cctgggacga ggggccccag gccggcggca
      901 agggggcccc gggctggcgc cacggggctg cgggccaccg tccgggcgaa ggtctgcagc
      961 gcctcgtact tggagcgcag cgcgtcgagc tccagcttca tgctggcgtt ctctgaggcc
     1021 agcttctcca cctcctgctg cagctccgcc ttctgcttct ccagctcctc cttctgcgtc
     1081 acccgcttca cgcggcagct ggcagcgtag ccgcggttct tgagcgtgcg ccggcgctgc
     1141 ttcagctgga cgatctcctc cttggacagg ccccgcaggt gctggttcag ctcccgcacc
     1201 gacatggtca ccagctcctc atccgtcagg ctggtgccat tctcacccgg ctcccgcttc
     1261 accttcaagg ccttgtttcc tttattgggg gtcgtcatgg cgatcgctag ctgcaaaaag
     1321 aacaagtagc ttgtattcta tagtgtcacc taaatgggcg atctgacggt tcactaaacc
     1381 agctctgctt atataggcct ccctggcgcg ccactagtta ttaatagtaa tcaattacgg
     1441 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     1501 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     1561 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     1621 cccacttggc agtacatcaa gtgtatcata tgccaagtcc gccccctatt gacgtcaatg
     1681 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttacgggac tttcctactt
     1741 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
     1801 ccaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     1861 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaataacc
     1921 ccgccccgtt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     1981 ctggtttagt gaaccgtcag atcactagaa gctttattgc ggtagtttat cacagttaaa
     2041 ttgctaacgc agtcagtgct tctgacacaa cagtctcgaa cttaagctgc agaagttggt
     2101 cgtgaggcac tgggcaggta agtatcaagg ttacaagaca ggtttaagga gaccaataga
     2161 aactgggctt gtcgagacag agaagactct tgcgtttctg ataggcacct attggtctta
     2221 ctgacatcca ctttgccttt ctctccacag gtgtccactc ccagttcaat tacagctctt
     2281 aaggctagag tattaatacg actcactata gggctagcga tcgccatgat ggacttggag
     2341 ctgccgccgc cgggactccc gtcccagcag gacatggatt tgattgacat actttggagg
     2401 caagatatag atcttggagt aagtcgagaa gtatttgact tcagtcagcg acggaaagag
     2461 tatgagctgg aaaaacagaa aaaacttgaa aaggaaagac aagaacaact ccaaaaggag
     2521 caagagaaag cctttttcgc tcagttacaa ctagatgaag agacaggtga atttctccca
     2581 attcagccag cccagcacat ccagtcagaa accagtggat ctgccaacta ctcccaggtt
     2641 gcccacattc ccaaatcaga tgctttgtac tttgatgact gcatgcagct tttggcgcag
     2701 acattcccgt ttgtagatga caatgaggtt tcttcggcta cgtttcagtc acttgttcct
     2761 gatattcccg gtcacatcga gagcccagtc ttcattgcta ctaatcaggc tcagtcacct
     2821 gaaacttctg ttgctcaggt agcccctgtt gatttagacg gtatgcaaca ggacattgag
     2881 caagtttggg aggagctatt atccattcct gagttacagt gtcttaatat tgaaaatgac
     2941 aagctggttg agactaccat ggttccaagt ccagaagcca aactgacaga agttgacaat
     3001 tatcattttt actcatctat accctcaatg gaaaaagaag taggtaactg tagtccacat
     3061 tttcttaatg cttttgagga ttccttcagc agcatcctct ccacagaaga ccccaaccag
     3121 ttgacagtga actcattaaa ttcagatgcc acagtcaaca cagattttgg tgatgaattt
     3181 tattctgctt tcatagctga gcccagtatc agcaacagca tgccctcacc tgctacttta
     3241 agccattcac tctctgaact tctaaatggg cccattgatg tttctgatct atcactttgc
     3301 aaagctttca accaaaacca ccctgaaagc acagcagaat tcaatgattc tgactccggc
     3361 atttcactaa acacaagtcc cagtgtggca tcaccagaac actcagtgga atcttccagc
     3421 tatggagaca cactacttgg cctcagtgat tctgaagtgg aagagctaga tagtgcccct
     3481 ggaagtgtca aacagaatgg tcctaaaaca ccagtacatt cttctgggga tatggtacaa
     3541 cccttgtcac catctcaggg gcagagcact cacgtgcatg atgcccaatg tgagaacaca
     3601 ccagagaaag aattgcctgt aagtcctggt catcggaaaa ccccattcac aaaagacaaa
     3661 cattcaagcc gcttggaggc tcatctcaca agagatgaac ttagggcaaa agctctccat
     3721 atcccattcc ctgtagaaaa aatcattaac ctccctgttg ttgacttcaa cgaaatgatg
     3781 tccaaagagc agttcaatga agctcaactt gcattaattc gggatatacg taggaggggt
     3841 aagaataaag tggctgctca gaattgcaga aaaagaaaac tggaaaatat agtagaacta
     3901 gagcaagatt tagatcattt gaaagatgaa aaagaaaaat tgctcaaaga aaaaggagaa
     3961 aatgacaaaa gccttcacct actgaaaaaa caactcagca ccttatatct cgaagttttc
     4021 agcatgctac gtgatgaaga tggaaaacct tattctccta gtgaatactc cctgcagcaa
     4081 acaagagatg gcaatgtttt ccttgttccc aaaagtaaga agccagatgt taagaaaaac
     4141 gtttctcaag gcagttcagg tggtggcggg agcggaggtg gaggctcgag cggtgtgacc
     4201 ggctaccggc tgttcgagga gattctgtaa ggccgcgact ctagagtcga cctgcaggca
     4261 tgcaagctga tttggctgct gccaccgctg agcaaggccg cttcgagcag acatgataag
     4321 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     4381 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     4441 caacaacaat tgcattcatt ttatgtttca ggttcagggg gagatgtggg aggttttttt
     4501 aagcaagtaa aacctctaca aatgtggtaa aatcttaatt aagaattcta atataacttc
     4561 gtatagcata cattatacga agttatggat ccactgttga caattaatca tcggcatagt
     4621 atatcggcat agtataatac gacaaggtga ggaactaaac ccaggaggca gatcatgatt
     4681 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     4741 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     4801 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
     4861 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     4921 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     4981 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     5041 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     5101 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
     5161 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
     5221 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     5281 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     5341 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     5401 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     5461 ttcttctgat tcgaaatgac cgaccaagcg acgcccaacc ggtatcagct cactcaaagg
     5521 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     5581 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     5641 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5701 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5761 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5821 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5881 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5941 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     6001 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     6061 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     6121 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     6181 agcagcagat tacgcgcaga aaaaaaggat ttcaagaaga tcctttgatc ttttctacgg
     6241 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     6301 aaaggatctt cacctagatc cttttatagt ccggaatcgt cacacaaaaa accaacacac
     6361 agatgtaatg aaaataaaga tattttattg cggccatcgt gatggctaga gtttagccct
     6421 cccacacata accagagggc agcaattcac gaatcccaac tgccgtcggc tgtccatcac
     6481 tgtccttcac tatggctttg atcccaggat gcagatcgag aagcacctgt cggcaccgtc
     6541 cgcaggggct caagatgccc ctgttctcat ttccgatcgc gacgatacaa gtcaggttgc
     6601 cagctgccgc agcagcagca gtgcccagca ccacgagttc tgcacaaggt cccccagtaa
     6661 aatgatatac attgacacca gtgaagatgc ggccgtcgct agagagagct gcgctggcga
     6721 cgctgtagtc ttcagagatg gggatgctgt tgattgtagc cgttgctctt tcaatgaggg
     6781 tggattcttc ttgagacaaa ggcttggcca tggtggcgct agtgtcagaa gaatcgagct
     6841 ttttgcaaaa gcctaggcct ccaaaaaagc ctcctcacta cttctggaat agctcagagg
     6901 ccgaggcggc ctcggcctct gcataaataa aaaaaattag tcagccatgg ggcggagaat
     6961 gggcggaact gggcggagtt aggggcggga tgggcggagt taggggcggg actatggttg
     7021 ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc
     7081 cacacctggt tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc
     7141 ctggggactt tccacaccct aactgacaca cattccacag ctggttcttt ccgcctcaga
     7201 aggtacctaa ccaagttcct ctttcagagg ttatttcagg ccatggtgct gcgc
//
