LOCUS       sequence-272407-        5673 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 5673)
  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 5673)
  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
FEATURES             Location/Qualifiers
     source          1..5673
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     3..27
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             39..104
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    complement(1961..2060)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2172..2191)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(2250..2269)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(2843..3376)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3565..3570,3571..3587,3588..3593))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      4250..4838
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4739..4758
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      5003..5089
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5181..5208
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     5216..5239
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    5258..5357
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(5389..5673,1..30)
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    5397..5415
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5433..5451
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5468..5486
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5504..5522
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5540..5558
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5575..5593
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5611..5629
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
ORIGIN
        1 agagctcgtt tagtgaaccg tcagatcgcc actagatgga ctacaaagac catgacggtg
       61 attataaaga tcatgacatc gattacaagg atgacgatga caagactagt atggacttgg
      121 agctgccgcc gccgggactc ccgtcccagc aggacatgga tttgattgac atactttgga
      181 ggcaagatat agatcttgga gtaagtcgag aagtatttga cttcagtcag cgacggaaag
      241 agtatgagct ggaaaaacag aaaaaacttg aaaaggaaag acaagaacaa ctccaaaagg
      301 agcaagagaa agcctttttc gctcagttac aactagatga agagacaggt gaatttctcc
      361 caattcagcc agcccagcac atccagtcag aaaccagtgg atctgccaac tactcccagg
      421 ttgcccacat tcccaaatca gatgctttgt actttgatga ctgcatgcag cttttggcgc
      481 agacattccc gtttgtagat gacaatgagg tttcttcggc tacgtttcag tcacttgttc
      541 ctgatattcc cggtcacatc gagagcccag tcttcattgc tactaatcag gctcagtcac
      601 ctgaaacttc tgttgctcag gtagcccctg ttgatttaga cggtatgcaa caggacattg
      661 agcaagtttg ggaggagcta ttatccattc ctgagttaca gtgtcttaat attgaaaatg
      721 acaagctggt tgagactacc atggttccaa gtccagaagc caaactgaca gaagttgaca
      781 attatcattt ttactcatct ataccctcaa tggaaaaaga agtaggtaac tgtagtccac
      841 attttcttaa tgcttttgag gattccttca gcagcatcct ctccacagaa gaccccaacc
      901 agttgacagt gaactcatta aattcagatg ccacagtcaa cacagatttt ggtgatgaat
      961 tttattctgc tttcatagct gagcccagta tcagcaacag catgccctca cctgctactt
     1021 taagccattc actctctgaa cttctaaatg ggcccattga tgtttctgat ctatcacttt
     1081 gcaaagcttt caaccaaaac caccctgaaa gcacagcaga attcaatgat tctgactccg
     1141 gcatttcact aaacacaagt cccagtgtgg catcaccaga acactcagtg gaatcttcca
     1201 gctatggaga cacactactt ggcctcagtg attctgaagt ggaagagcta gatagtgccc
     1261 ctggaagtgt caaacagaat ggtcctaaaa caccagtaca ttcttctggg gatatggtac
     1321 aacccttgtc accatctcag gggcagagca ctcacgtgca tgatgcccaa tgtgagaaca
     1381 caccagagaa agaattgcct gtaagtcctg gtcatcggaa aaccccattc acaaaagaca
     1441 aacattcaag ccgcttggag gctcatctca caagagatga acttagggca aaagctctcc
     1501 atatcccatt ccctgtagaa aaaatcatta acctccctgt tgttgacttc aacgaaatga
     1561 tgtccaaaga gcagttcaat gaagctcaac ttgcattaat tcgggatata cgtaggaggg
     1621 gtaagaataa agtggctgct cagaattgca gaaaaagaaa actggaaaat atagtagaac
     1681 tagagcaaga tttagatcat ttgaaagatg aaaaagaaaa attgctcaaa gaaaaaggag
     1741 aaaatgacaa aagccttcac ctactgaaaa aacaactcag caccttatat ctcgaagttt
     1801 tcagcatgct acgtgatgaa gatggaaaac cttattctcc tagtgaatac tccctgcagc
     1861 aaacaagaga tggcaatgtt ttccttgttc ccaaaagtaa gaagccagat gttaagaaaa
     1921 actagcaatt ggaagactaa tgcgttttcg agatatctag acccagcttt cttgtacaaa
     1981 gttggcatta taagaaagca ttgcttatca atttgttgca acgaacaggt cactatcagt
     2041 caaaataaaa tcattatttg ccatccagct gcagctctgg cccgtgtctc aaaatctctg
     2101 atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt ctgcttacat
     2161 aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtcga ggccgcgatt
     2221 aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca
     2281 atcaggtgcg acaatctatc gcttgtatgg gaagcccgat gcgccagagt tgtttctgaa
     2341 acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct
     2401 gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg
     2461 gttactcacc actgcgatcc ccggaaaaac agcattccag gtattagaag aatatcctga
     2521 ttcaggtgaa aatattgttg atgcgctggc agtgtccctg cgccggttgc attcgattcc
     2581 tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg
     2641 aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt
     2701 tgaacaagtc tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac
     2761 tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattag gccgggaagc
     2821 cgatctcggc ttgaacgaat tgttaggtgg cggtacttgg gtcgatatca aagtgcatca
     2881 cttcttcccg tatgcccaac tttgtataga gagccactgc gggatcgtca ccgtaatctg
     2941 cttgcacgta gatcacataa gcaccaagcg cgttggcctc atgcttgagg agattgatga
     3001 gcgcggtggc aatgccctgc ctccggtgct cgccggagac tgcgagatca tagatataga
     3061 tctcactacg cggctgctca aacttgggca gaacgtaagc cgcgagagcg ccaacaaccg
     3121 cttcttggtc gaaggcagca agcgcgatga atgtcttact acggagcaag ttcccgaggt
     3181 aatcggagtc cggctgatgt tgggagtagg tggctacgtc tccgaactca cgaccgaaaa
     3241 gatcaagagc agcccgcatg gatttgactt ggtcagggcc gagcctacat gtgcgaatga
     3301 tgcccatact tgagccacct aactttgttt tagggcgact gccctgctgc gtaacatcgt
     3361 tgctgctgcg taacatcgtt gctgctccat aacatcaaac atcgacccac ggcgtaacgc
     3421 gcttgctgct tggatgcccg aggcatagac tgtacaaaaa aacagtcata acaagccatg
     3481 aaaaccgcca ctgcgccgtt accaccgctg cgttcggtca aggttctgga ccagttgcgt
     3541 gagcgcatac gctacttgca ttacagttta cgaaccgaac aggcttatgt caactgggtt
     3601 cgtgccttca tccgtttcca cggtgtgcgt cacccggcaa ccttgggcag cagcgaagtc
     3661 gaggcatttc tgtcctggct ggcgaacgag cgcaaggttt cggtctccac gcatcgtcag
     3721 gcattggcgg ccttgctgtt cttctacggc aaggtgctgt gcacggatct gccctggctt
     3781 caggagatcg gaagacctcg gccgtcgcgg cgcttgccgg tggtgctgac cccggatgaa
     3841 gtggttcgca tcctcggttt tctggaaggc gagcatcgtt tgttcgccca ggactctagc
     3901 tatagttcta gtggttggct actaataggt tgtattgatg ttggacgagt cggaatcgca
     3961 gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc tccttcatta
     4021 cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa attgcagttt
     4081 catttgatgc tcgatgagtt tttctaatca gaattggtta attggttgta acactggcag
     4141 agcattacgc tgacttgacg ggacggcgca agctcatgac caaaatccct taacgtgagt
     4201 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     4261 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     4321 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     4381 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     4441 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     4501 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     4561 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     4621 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     4681 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     4741 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     4801 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     4861 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     4921 attctgtgga taaccgtatt accgctagca tggatctcgg ggacgtctaa ctactaagcg
     4981 agagtaggga actgccaggc atcaaataaa acgaaaggct cagtcggaag actgggcctt
     5041 tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc cgccgggagc
     5101 ggatttgaac gttgtgaagc aacggcccgg agggtggcgg gcaggacgcc cgccataaac
     5161 tgccaggcat caaactaagc agaaggccat cctgacggat ggcctttttg cgtttctaca
     5221 aactcttcct gttagttagt tacttaagct cgggccccaa ataatgattt tattttgact
     5281 gatagtgacc tgttcgttgc aacaaattga taagcaatgc ttttttataa tgccaacttt
     5341 gtacaaaaaa gcaggcttta aaggaaccaa ttcagtcgac tggatcccga gtttactccc
     5401 tatcagtgat agagaacgta tgtcgagttt actccctatc agtgatagag aacgatgtcg
     5461 agtttactcc ctatcagtga tagagaacgt atgtcgagtt tactccctat cagtgataga
     5521 gaacgtatgt cgagtttact ccctatcagt gatagagaac gtatgtcgag tttatcccta
     5581 tcagtgatag agaacgtatg tcgagtttac tccctatcag tgatagagaa cgtatgtcga
     5641 ggtaggcgtg tacggtggga ggcctatata agc
//
