LOCUS       sequence-474708-        7455 bp    DNA     circular SYN 05-JUN-2025
DEFINITION  Express NRF2- SmBiT 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 7455)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 7455)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 5, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: NRF2-SmBiT Fusion Vector
FEATURES             Location/Qualifiers
     source          1..7455
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    98..146
                     /note="synthetic polyadenylation signal"
     rep_origin      272..860
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     761..780
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1018..1812)
                     /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(1129..1148)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1739..1758
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1844..1891)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     protein_bind    1901..1934
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     polyA_signal    2049..2170
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     2140..2159
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4174..4193)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4175..4193)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4235..4254)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          4237..4369
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        complement(4505..4708)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4534..4554)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4709..5012
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     polyA_signal    5245..5326
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5428..5448)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6904..7100
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6951..7086
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      7125..7134
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(7131..7455,1..74)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
ORIGIN
        1 aggacagtga tggacagccg acggcagttg ggattcgtga attgctgccc tctggttatg
       61 tgtgggaggg ctaaactcta gccatcacga tggccgcaat aaaatatctt tattttcatt
      121 acatctgtgt gttggttttt tgtgtgacga ttccggacta taaaaggatc taggtgaaga
      181 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
      241 cagaccccgt agaaaagatc aaaggatctt cttgaaatcc tttttttctg cgcgtaatct
      301 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
      361 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
      421 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
      481 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
      541 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
      601 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
      661 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
      721 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
      781 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
      841 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
      901 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
      961 ttaccgcctt tgagtgagct gataccggtt gggcgtcgct tggtcggtca tttcgaatca
     1021 gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc
     1081 gtaaagcacg aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt
     1141 agccaacgct atgtcctgat agcggtccgc cacacccagc cggccacagt cgatgaatcc
     1201 agaaaagcgg ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac
     1261 gagatcctcg ccgtcgggca tgcgcgcctt gagcctggcg aacagttcgg ctggcgcgag
     1321 cccctgatgc tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg
     1381 tgctcgctcg atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt
     1441 atgcagccgc cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga
     1501 tgacaggaga tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt
     1561 gacaacgtcg agcacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc
     1621 tgcctcgtcc tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg
     1681 gcgcccctgc gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc
     1741 ccagtcatag ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc
     1801 ttgttcaatc atgatctgcc tcctgggttt agttcctcac cttgtcgtat tatactatgc
     1861 cgatatacta tgccgatgat taattgtcaa cagtggatcc ataacttcgt ataatgtatg
     1921 ctatacgaag ttatattaga attcttaatt aagattttac cacatttgta gaggttttac
     1981 ttgcttaaaa aaacctccca catctccccc tgaacctgaa acataaaatg aatgcaattg
     2041 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2101 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     2161 atgtatctta tcatgtctgc tcgaagcggc cttgctcagc ggtggcagca gccaaatcag
     2221 cttgcatgcc tgcaggtcga ctctagagtc gcggccttac agaatctcct cgaacagccg
     2281 gtagccggtc acaccgctcg agcctccacc tccgctcccg ccaccacctg aactgccttg
     2341 agaaacgttt ttcttaacat ctggcttctt acttttggga acaaggaaaa cattgccatc
     2401 tcttgtttgc tgcagggagt attcactagg agaataaggt tttccatctt catcacgtag
     2461 catgctgaaa acttcgagat ataaggtgct gagttgtttt ttcagtaggt gaaggctttt
     2521 gtcattttct cctttttctt tgagcaattt ttctttttca tctttcaaat gatctaaatc
     2581 ttgctctagt tctactatat tttccagttt tctttttctg caattctgag cagccacttt
     2641 attcttaccc ctcctacgta tatcccgaat taatgcaagt tgagcttcat tgaactgctc
     2701 tttggacatc atttcgttga agtcaacaac agggaggtta atgatttttt ctacagggaa
     2761 tgggatatgg agagcttttg ccctaagttc atctcttgtg agatgagcct ccaagcggct
     2821 tgaatgtttg tcttttgtga atggggtttt ccgatgacca ggacttacag gcaattcttt
     2881 ctctggtgtg ttctcacatt gggcatcatg cacgtgagtg ctctgcccct gagatggtga
     2941 caagggttgt accatatccc cagaagaatg tactggtgtt ttaggaccat tctgtttgac
     3001 acttccaggg gcactatcta gctcttccac ttcagaatca ctgaggccaa gtagtgtgtc
     3061 tccatagctg gaagattcca ctgagtgttc tggtgatgcc acactgggac ttgtgtttag
     3121 tgaaatgccg gagtcagaat cattgaattc tgctgtgctt tcagggtggt tttggttgaa
     3181 agctttgcaa agtgatagat cagaaacatc aatgggccca tttagaagtt cagagagtga
     3241 atggcttaaa gtagcaggtg agggcatgct gttgctgata ctgggctcag ctatgaaagc
     3301 agaataaaat tcatcaccaa aatctgtgtt gactgtggca tctgaattta atgagttcac
     3361 tgtcaactgg ttggggtctt ctgtggagag gatgctgctg aaggaatcct caaaagcatt
     3421 aagaaaatgt ggactacagt tacctacttc tttttccatt gagggtatag atgagtaaaa
     3481 atgataattg tcaacttctg tcagtttggc ttctggactt ggaaccatgg tagtctcaac
     3541 cagcttgtca ttttcaatat taagacactg taactcagga atggataata gctcctccca
     3601 aacttgctca atgtcctgtt gcataccgtc taaatcaaca ggggctacct gagcaacaga
     3661 agtttcaggt gactgagcct gattagtagc aatgaagact gggctctcga tgtgaccggg
     3721 aatatcagga acaagtgact gaaacgtagc cgaagaaacc tcattgtcat ctacaaacgg
     3781 gaatgtctgc gccaaaagct gcatgcagtc atcaaagtac aaagcatctg atttgggaat
     3841 gtgggcaacc tgggagtagt tggcagatcc actggtttct gactggatgt gctgggctgg
     3901 ctgaattggg agaaattcac ctgtctcttc atctagttgt aactgagcga aaaaggcttt
     3961 ctcttgctcc ttttggagtt gttcttgtct ttccttttca agttttttct gtttttccag
     4021 ctcatactct ttccgtcgct gactgaagtc aaatacttct cgacttactc caagatctat
     4081 atcttgcctc caaagtatgt caatcaaatc catgtcctgc tgggacggga gtcccggcgg
     4141 cggcagctcc aagtccatca tggcgatcgc tagccctata gtgagtcgta ttaatactct
     4201 agccttaaga gctgtaattg aactgggagt ggacacctgt ggagagaaag gcaaagtgga
     4261 tgtcagtaag accaataggt gcctatcaga aacgcaagag tcttctctgt ctcgacaagc
     4321 ccagtttcta ttggtctcct taaacctgtc ttgtaacctt gatacttacc tgcccagtgc
     4381 ctcacgacca acttctgcag cttaagttcg agactgttgt gtcagaagca ctgactgcgt
     4441 tagcaattta actgtgataa actaccgcaa taaagcttct agtgatctga cggttcacta
     4501 aaccagctct gcttatatag acctcccacc gtacacgcct accgcccatt tgcgtcaacg
     4561 gggcggggtt attacgacat tttggaaagt cccgttgatt ttggtgccaa aacaaactcc
     4621 cattgacgtc aatggggtgg agacttggaa atccccgtga gtcaaaccgc tatccacgcc
     4681 cattggtgta ctgccaaaac cgcatcacca tggtaatagc gatgactaat acgtagatgt
     4741 actgccaagt aggaaagtcc cgtaaggtca tgtactgggc ataatgccag gcgggccatt
     4801 taccgtcatt gacgtcaata gggggcggac ttggcatatg atacacttga tgtactgcca
     4861 agtgggcagt ttaccgtaaa tactccaccc attgacgtca atggaaagtc cctattggcg
     4921 ttactatggg aacatacgtc attattgacg tcaatgggcg ggggtcgttg ggcggtcagc
     4981 caggcgggcc atttaccgta agttatgtaa cgcggaactc catatatggg ctatgaacta
     5041 atgaccccgt aattgattac tattaataac tagtggcgcg ccataatcct aggctctcgt
     5101 acgaagtcag ctgataacgt cgacgtattg cggccgcgga tcataatcag ccataccaca
     5161 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     5221 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     5281 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgccgac gtcttaatta
     5341 acgcgtcttg cgcagcacca tggcctgaaa taacctctga aagaggaact tggttaggta
     5401 ccttctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     5461 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     5521 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcactgag agatcccctc
     5581 ataatttccc caaagcgtaa ccatgtgtga ataaattttg agctagtagg gttgcagcca
     5641 cgagtaagtc ttcccttgtt attgtgtagc cagaatgccg caaaacttcc atgcctaagc
     5701 gaactgttga gagtacgttt cgatttctga ctgtgttagc ctggaagtgc ttgtcccaac
     5761 cttgtttctg agcatgaacg cccgcaagcc aacatgttag ttgaagcatc agggcgatta
     5821 gcagcatgat atcaaaacgc tctgagctgc tcgttcggct atggcgtagg cctagtccgt
     5881 aggcaggact tttcaagtct cggaaggttt cttcaatctg cattcgcttc gaatagatat
     5941 taacaagttg tttgggtgtt cgaatttcaa caggtaagtt agttgctaga acccatggct
     6001 cctttgccga cgctgagtag attttaggtg acgggtggtg acaatgagtc cgtgtcgagc
     6061 gctgattttt tcggccttta gagcgagatt tatacaatag aatttggcat gagattggat
     6121 tgcttttagt cagcctctta tagcctaaag tctttgagtg actagatgac atatcatgta
     6181 agttgctgat aggtttccag ttttccgctc ctaggtctgc atattgtact tttcctctta
     6241 ctcgacttaa ccagtaccaa cccagcttct caacggattt ataccatggc actttaaagc
     6301 cagcatcact gacaatgagc ggtgtggtgt tactcggtag aatgctcgca aggtcggcta
     6361 gaaattggtc atgagctttc tttgaacatt gctctgaaag cgggaacgct ttctcataaa
     6421 gagtaacaga acgaccgtgt agtgcgactg aagctcgcaa taccataagt cgtttttgct
     6481 cacgaatatc agaccagtca acaagtacaa tgggcatcgt attgcccgaa cagataaagc
     6541 tagcatgcca acggtataca gcgagtcgct ctttgtggag gtgacgatta cctaacaatc
     6601 ggtcgattcg tttgatgtta tgttttgttc tcgctttggt tggcaggtta cggccaagtt
     6661 cggtaagagt gagagtttta cagtcaagta atgcgtggca agccaacgtt aagctgttga
     6721 gtcgttttaa gtgtaattcg gggcagaatt ggtaaagaga gtcgtgtaaa atatcgagtt
     6781 cgcacatctt gttgtctgat tattgatttt tcgcgaaacc atttgatcat atgacaagat
     6841 gtgtatccac cttaacttaa tgatttttac caaaatcatt aggggattca tcagtgcaaa
     6901 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     6961 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     7021 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
     7081 gaggcctagg cttttgcaaa aagctcgatt cttctgacac tagcgccacc atggccaagc
     7141 ctttgtctca agaagaatcc accctcattg aaagagcaac ggctacaatc aacagcatcc
     7201 ccatctctga agactacagc gtcgccagcg cagctctctc tagcgacggc cgcatcttca
     7261 ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc agaactcgtg gtgctgggca
     7321 ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt cgcgatcgga aatgagaaca
     7381 ggggcatctt gagcccctgc ggacggtgcc gacaggtgct tctcgatctg catcctggga
     7441 tcaaagccat agtga
//
