LOCUS       sequence-272397-        5603 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used as a donor vector containing N-term 3xFLAG to clone into pSLIK.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5603)
  AUTHORS   Pereira EJ, Burns JS, Lee CY, Marohl T, Calderon D, Wang L, Atkins 
            KA, Wang CC, Janes KA
  TITLE     Sporadic activation of an oxidative stress-dependent NRF2-p53 
            signaling network in breast epithelial spheroids and 
            premalignancies.
  JOURNAL   Sci Signal. 2020 Apr 14;13(627). pii: 13/627/eaba4200. doi: 
            10.1126/scisignal.aba4200.
  PUBMED    32291314
REFERENCE   2  (bases 1 to 5603)
  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: "Sci 
            Signal."; date: "2020-04-14"; volume: "13(627). pii: 
            13/627/eaba4200. doi"; pages: " 10.1126/scisignal.aba4200"
COMMENT     Sequence Label: pEN_TTmiRC2_3xflag_NRF2(RR)
FEATURES             Location/Qualifiers
     source          1..5603
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    18..36
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    54..72
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    90..108
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    126..144
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        156..194
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     191..215
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             227..292
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    complement(2149..2248)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2360..2379)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(2438..2457)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(3031..3564)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3753..3758,3759..3775,3776..3781))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      4438..5026
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4927..4946
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      5191..5277
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5369..5396
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     5404..5427
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    5446..5545
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    5585..5603
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
ORIGIN
        1 acgtatgtcg agtttactcc ctatcagtga tagagaacgn atgtcgagtt tactccctat
       61 cagtgataga gaacgtatgt cgagtttact ccctatcagt gatagagaac gtatgtcgag
      121 tttactccct atcagtgata gagaacgtat gtcgaggtag gcgtgtacgg tgggaggcct
      181 atataagcag agctcgttta gtgaaccgtc agatcgccac tagatggact acaaagacca
      241 tgacggtgat tataaagatc atgacatcga ttacaaggat gacgatgaca agactagtat
      301 ggacttggag ctgccgccgc cgggactccc gtcccagcag gacatggatt tgattgacat
      361 actttggagg caagatatag atcttggagt aagtcgagaa gtatttgact tcagtcagcg
      421 acggaaagag tatgagctgg aaaaacagaa aaaacttgaa aaggaaagac aagaacaact
      481 ccaaaaggag caagagaaag cctttttcgc tcagttacaa ctagatgaag agacaggtga
      541 atttctccca attcagccag cccagcacat ccagtcagaa accagtggat ctgccaacta
      601 ctcccaggtt gcccacattc ccaaatcaga tgctttgtac tttgatgact gcatgcagct
      661 tttggcgcag acattcccgt ttgtagatga caatgaggtt tcttcggcta cgtttcagtc
      721 acttgttcct gatattcccg gtcacatcga gagcccagtc ttcattgcta ctaatcaggc
      781 tcagtcacct gaaacttctg ttgctcaggt agcccctgtt gatttagacg gtatgcaaca
      841 ggacattgag caagtttggg aggagctatt atccattcct gagttacagt gtcttaatat
      901 tgaaaatgac aagctggttg agactaccat ggttccaagt ccagaagcca aactgacaga
      961 agttgacaat tatcattttt actcatctat accctcaatg gaaaaagaag taggtaactg
     1021 tagtccacat tttcttaatg cttttgagga ttccttcagc agcatcctct ccacagaaga
     1081 ccccaaccag ttgacagtga actcattaaa ttcagatgcc acagtcaaca cagattttgg
     1141 tgatgaattt tattctgctt tcatagctga gcccagtatc agcaacagca tgccctcacc
     1201 tgctacttta agccattcac tctctgaact tctaaatggg cccattgatg tttctgatct
     1261 atcactttgc aaagctttca accaaaacca ccctgaaagc acagcagaat tcaatgattc
     1321 tgactccggc atttcactaa acacaagtcc cagtgtggca tcaccagaac actcagtgga
     1381 atcttccagc tatggagaca cactacttgg cctcagtgat tctgaagtgg aagagctaga
     1441 tagtgcccct ggaagtgtca aacagaatgg tcctaaaaca ccagtacatt cttctgggga
     1501 tatggtacaa cccttgtcac catctcaggg gcagagcact cacgtgcatg atgcccaatg
     1561 tgagaacaca ccagagaaag aattgcctgt aagtcctggt catcggaaaa ccccattcac
     1621 aaaagacaaa cattcaagcc gcttggaggc tcatctcaca agagatgaac ttagggcaaa
     1681 agctctccat atcccattcc ctgtagaaaa aatcattaac ctccctgttg ttgacttcaa
     1741 cgaaatgatg tccaaagagc agttcaatga agctcaactt gcattaattc gggatatacg
     1801 taggaggggt aagaataaag tggctgctca gaattgcaga aaaagaaaac tggaaaatat
     1861 agtagaacta gagcaagatt tcgttcgttt gaaagatgaa aaagaaaaat tgctcaaaga
     1921 aaaaggagaa aatgacaaaa gccttcacct actgaaaaaa caactcagca ccttatatct
     1981 cgaagttttc agcatgctac gtgatgaaga tggaaaacct tattctccta gtgaatactc
     2041 cctgcagcaa acaagagatg gcaatgtttt ccttgttccc aaaagtaaga agccagatgt
     2101 taagaaaaac tagcaattgg aagactaatg cgttttcgag atatctagac ccagctttct
     2161 tgtacaaagt tggcattata agaaagcatt gcttatcaat ttgttgcaac gaacaggtca
     2221 ctatcagtca aaataaaatc attatttgcc atccagctgc agctctggcc cgtgtctcaa
     2281 aatctctgat gttacattgc acaagataaa aatatatcat catgaacaat aaaactgtct
     2341 gcttacataa acagtaatac aaggggtgtt atgagccata ttcaacggga aacgtcgagg
     2401 ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat
     2461 gtcgggcaat caggtgcgac aatctatcgc ttgtatggga agcccgatgc gccagagttg
     2521 tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta
     2581 aactggctga cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat
     2641 gatgcatggt tactcaccac tgcgatcccc ggaaaaacag cattccaggt attagaagaa
     2701 tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgtccctgcg ccggttgcat
     2761 tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg
     2821 caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc
     2881 tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca
     2941 gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaggc
     3001 cgggaagccg atctcggctt gaacgaattg ttaggtggcg gtacttgggt cgatatcaaa
     3061 gtgcatcact tcttcccgta tgcccaactt tgtatagaga gccactgcgg gatcgtcacc
     3121 gtaatctgct tgcacgtaga tcacataagc accaagcgcg ttggcctcat gcttgaggag
     3181 attgatgagc gcggtggcaa tgccctgcct ccggtgctcg ccggagactg cgagatcata
     3241 gatatagatc tcactacgcg gctgctcaaa cttgggcaga acgtaagccg cgagagcgcc
     3301 aacaaccgct tcttggtcga aggcagcaag cgcgatgaat gtcttactac ggagcaagtt
     3361 cccgaggtaa tcggagtccg gctgatgttg ggagtaggtg gctacgtctc cgaactcacg
     3421 accgaaaaga tcaagagcag cccgcatgga tttgacttgg tcagggccga gcctacatgt
     3481 gcgaatgatg cccatacttg agccacctaa ctttgtttta gggcgactgc cctgctgcgt
     3541 aacatcgttg ctgctgcgta acatcgttgc tgctccataa catcaaacat cgacccacgg
     3601 cgtaacgcgc ttgctgcttg gatgcccgag gcatagactg tacaaaaaaa cagtcataac
     3661 aagccatgaa aaccgccact gcgccgttac caccgctgcg ttcggtcaag gttctggacc
     3721 agttgcgtga gcgcatacgc tacttgcatt acagtttacg aaccgaacag gcttatgtca
     3781 actgggttcg tgccttcatc cgtttccacg gtgtgcgtca cccggcaacc ttgggcagca
     3841 gcgaagtcga ggcatttctg tcctggctgg cgaacgagcg caaggtttcg gtctccacgc
     3901 atcgtcaggc attggcggcc ttgctgttct tctacggcaa ggtgctgtgc acggatctgc
     3961 cctggcttca ggagatcgga agacctcggc cgtcgcggcg cttgccggtg gtgctgaccc
     4021 cggatgaagt ggttcgcatc ctcggttttc tggaaggcga gcatcgtttg ttcgcccagg
     4081 actctagcta tagttctagt ggttggctac taataggttg tattgatgtt ggacgagtcg
     4141 gaatcgcaga ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc
     4201 cttcattaca gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat
     4261 tgcagtttca tttgatgctc gatgagtttt tctaatcaga attggttaat tggttgtaac
     4321 actggcagag cattacgctg acttgacggg acggcgcaag ctcatgacca aaatccctta
     4381 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     4441 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     4501 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     4561 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     4621 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     4681 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     4741 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     4801 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     4861 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     4921 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     4981 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     5041 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     5101 atcccctgat tctgtggata accgtattac cgctagcatg gatctcgggg acgtctaact
     5161 actaagcgag agtagggaac tgccaggcat caaataaaac gaaaggctca gtcggaagac
     5221 tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg
     5281 ccgggagcgg atttgaacgt tgtgaagcaa cggcccggag ggtggcgggc aggacgcccg
     5341 ccataaactg ccaggcatca aactaagcag aaggccatcc tgacggatgg cctttttgcg
     5401 tttctacaaa ctcttcctgt tagttagtta cttaagctcg ggccccaaat aatgatttta
     5461 ttttgactga tagtgacctg ttcgttgcaa caaattgata agcaatgctt ttttataatg
     5521 ccaactttgt acaaaaaagc aggctttaaa ggaaccaatt cagtcgactg gatcccgagt
     5581 ttactcccta tcagtgatag aga
//
