LOCUS       sequence-272437-        5248 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used as a donor vector to clone into pSLIK.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5248)
  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 5248)
  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_BirA_CDKN1A
FEATURES             Location/Qualifiers
     source          1..5248
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        25..339
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    33..51
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    69..87
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    104..122
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    140..158
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    176..194
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    211..229
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    247..265
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     312..336
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(346..696,697..699,700..1308)
                     /codon_start=1
                     /product="promiscuous R118G mutant of the E. coli biotin 
                     protein ligase (Cronan, 2005)"
                     /label=BirA(R118G)
                     /note="also known as BirA* or BioID"
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGGRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             1315..1809
                     /codon_start=1
                     /gene="Homo sapiens CDKN1A"
                     /product="human cyclin-dependent kinase inhibitor 1A"
                     /label=p21
                     /note="mediates p53-dependent arrest in the G1 phase of the
                     cell cycle"
                     /translation="MSEPAGDVRQNPCGSKACRRLFGPVDSEQLRRDCDALMAGCIQEA
                     RERWNFDFVTETPLEGDFAWERVRGLGLPKLYLPTGPRRGRDELGGGRRPGTSPALLQG
                     TAEEDHVDLSLSCTLVPRSGEQAEGSPGGPGDSQGRKRRQTSMTDFYHSKRRLIFSKRK
                     P"
     protein_bind    complement(1845..1944)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2056..2075)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(2134..2153)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(2727..3260)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3449..3454,3455..3471,3472..3477))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      4134..4722
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4623..4642
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      4887..4973
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5065..5092
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     5100..5123
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    5142..5241
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
ORIGIN
        1 aaccaattca gtcgactgga tcccgagttt actccctatc agtgatagag aacgtatgtc
       61 gagtttactc cctatcagtg atagagaacg atgtcgagtt tactccctat cagtgataga
      121 gaacgtatgt cgagtttact ccctatcagt gatagagaac gtatgtcgag tttactccct
      181 atcagtgata gagaacgtat gtcgagttta tccctatcag tgatagagaa cgtatgtcga
      241 gtttactccc tatcagtgat agagaacgta tgtcgaggta ggcgtgtacg gtgggaggcc
      301 tatataagca gagctcgttt agtgaaccgt cagatcgcca ctagaatgaa ggacaacacc
      361 gtgcccctga agctgatcgc cctgctggcc aacggcgagt tccactctgg cgagcagctg
      421 ggagagaccc tgggaatgag cagagccgcc atcaacaagc acatccagac actgagagac
      481 tggggagtgg acgtgttcac cgtgcctggc aagggctaca gcctgcctga gcctatccag
      541 ctgctgaacg ccaagcagat cctgggacag ctggatggcg gaagcgtggc cgtgctgcct
      601 gtgatcgact ccaccaatca gtacctgctg gacagaatcg gagagctgaa gtccggcgac
      661 gcctgcatcg ccgagtacca gcaggctggc agaggaggca gaggacggaa gtggttcagc
      721 ccattcggag ccaacctgta cctgtccatg ttctggagac tggagcaggg acctgctgct
      781 gccatcggac tgagtctggt gatcggaatc gtgatggccg aggtgctgag aaagctggga
      841 gccgacaagg tgagagtgaa gtggcctaat gacctgtacc tccaggaccg caagctggct
      901 ggcatcctgg tggagctgac aggcaagaca ggcgatgccg ctcagatcgt gatcggagcc
      961 ggaatcaaca tggccatgag aagagtggag gagagcgtgg tgaaccaggg ctggatcacc
     1021 ctgcaggagg ctggcatcaa cctggaccgg aacaccctgg ccgccatgct gatcagagag
     1081 ctgagagccg ctctggagct gttcgagcag gagggactgg ctccttacct gagcagatgg
     1141 gagaagctgg acaacttcat caacagacct gtgaagctga tcatcggcga caaggaaatc
     1201 ttcggcatct ccagaggaat cgacaagcag ggagctctgc tgctggagca ggacggaatc
     1261 atcaagccct ggatgggcgg agaaatctcc ctgagaagcg cagagaagac tagtatgtca
     1321 gaaccggctg gggatgtccg tcagaaccca tgcggcagca aggcctgccg ccgcctcttc
     1381 ggcccagtgg acagcgagca gctgagacgc gactgtgatg cgctaatggc gggctgcatc
     1441 caggaggccc gtgagcgatg gaacttcgac tttgtcaccg agacaccact ggagggtgac
     1501 ttcgcctggg agcgtgtgcg gggccttggc ctgcccaagc tctaccttcc cacggggccc
     1561 cggcgaggcc gggatgagtt gggaggaggc aggcggcctg gcacctcacc tgctctgctg
     1621 caggggacag cagaggaaga ccatgtggac ctgtcactgt cttgtaccct tgtgcctcgc
     1681 tcaggggagc aggctgaagg gtccccaggt ggacctggag actctcaggg tcgaaaacgg
     1741 cggcagacca gcatgacaga tttctaccac tccaaacgcc ggctgatctt ctccaagagg
     1801 aagccctaac aattggaaga ctaatgcgtt ttcgagatat ctagacccag ctttcttgta
     1861 caaagttggc attataagaa agcattgctt atcaatttgt tgcaacgaac aggtcactat
     1921 cagtcaaaat aaaatcatta tttgccatcc agctgcagct ctggcccgtg tctcaaaatc
     1981 tctgatgtta cattgcacaa gataaaaata tatcatcatg aacaataaaa ctgtctgctt
     2041 acataaacag taatacaagg ggtgttatga gccatattca acgggaaacg tcgaggccgc
     2101 gattaaattc caacatggat gctgatttat atgggtataa atgggctcgc gataatgtcg
     2161 ggcaatcagg tgcgacaatc tatcgcttgt atgggaagcc cgatgcgcca gagttgtttc
     2221 tgaaacatgg caaaggtagc gttgccaatg atgttacaga tgagatggtc agactaaact
     2281 ggctgacgga atttatgcct cttccgacca tcaagcattt tatccgtact cctgatgatg
     2341 catggttact caccactgcg atccccggaa aaacagcatt ccaggtatta gaagaatatc
     2401 ctgattcagg tgaaaatatt gttgatgcgc tggcagtgtc cctgcgccgg ttgcattcga
     2461 ttcctgtttg taattgtcct tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat
     2521 cacgaatgaa taacggtttg gttgatgcga gtgattttga tgacgagcgt aatggctggc
     2581 ctgttgaaca agtctggaaa gaaatgcata aacttttgcc attctcaccg gattcagtcg
     2641 tcactcatgg tgatttctca cttgataacc ttatttttga cgaggggaaa ttaggccggg
     2701 aagccgatct cggcttgaac gaattgttag gtggcggtac ttgggtcgat atcaaagtgc
     2761 atcacttctt cccgtatgcc caactttgta tagagagcca ctgcgggatc gtcaccgtaa
     2821 tctgcttgca cgtagatcac ataagcacca agcgcgttgg cctcatgctt gaggagattg
     2881 atgagcgcgg tggcaatgcc ctgcctccgg tgctcgccgg agactgcgag atcatagata
     2941 tagatctcac tacgcggctg ctcaaacttg ggcagaacgt aagccgcgag agcgccaaca
     3001 accgcttctt ggtcgaaggc agcaagcgcg atgaatgtct tactacggag caagttcccg
     3061 aggtaatcgg agtccggctg atgttgggag taggtggcta cgtctccgaa ctcacgaccg
     3121 aaaagatcaa gagcagcccg catggatttg acttggtcag ggccgagcct acatgtgcga
     3181 atgatgccca tacttgagcc acctaacttt gttttagggc gactgccctg ctgcgtaaca
     3241 tcgttgctgc tgcgtaacat cgttgctgct ccataacatc aaacatcgac ccacggcgta
     3301 acgcgcttgc tgcttggatg cccgaggcat agactgtaca aaaaaacagt cataacaagc
     3361 catgaaaacc gccactgcgc cgttaccacc gctgcgttcg gtcaaggttc tggaccagtt
     3421 gcgtgagcgc atacgctact tgcattacag tttacgaacc gaacaggctt atgtcaactg
     3481 ggttcgtgcc ttcatccgtt tccacggtgt gcgtcacccg gcaaccttgg gcagcagcga
     3541 agtcgaggca tttctgtcct ggctggcgaa cgagcgcaag gtttcggtct ccacgcatcg
     3601 tcaggcattg gcggccttgc tgttcttcta cggcaaggtg ctgtgcacgg atctgccctg
     3661 gcttcaggag atcggaagac ctcggccgtc gcggcgcttg ccggtggtgc tgaccccgga
     3721 tgaagtggtt cgcatcctcg gttttctgga aggcgagcat cgtttgttcg cccaggactc
     3781 tagctatagt tctagtggtt ggctactaat aggttgtatt gatgttggac gagtcggaat
     3841 cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc
     3901 attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca
     3961 gtttcatttg atgctcgatg agtttttcta atcagaattg gttaattggt tgtaacactg
     4021 gcagagcatt acgctgactt gacgggacgg cgcaagctca tgaccaaaat cccttaacgt
     4081 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4141 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     4201 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     4261 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     4321 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     4381 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     4441 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     4501 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     4561 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     4621 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     4681 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     4741 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     4801 cctgattctg tggataaccg tattaccgct agcatggatc tcggggacgt ctaactacta
     4861 agcgagagta gggaactgcc aggcatcaaa taaaacgaaa ggctcagtcg gaagactggg
     4921 cctttcgttt tatctgttgt ttgtcggtga acgctctcct gagtaggaca aatccgccgg
     4981 gagcggattt gaacgttgtg aagcaacggc ccggagggtg gcgggcagga cgcccgccat
     5041 aaactgccag gcatcaaact aagcagaagg ccatcctgac ggatggcctt tttgcgtttc
     5101 tacaaactct tcctgttagt tagttactta agctcgggcc ccaaataatg attttatttt
     5161 gactgatagt gacctgttcg ttgcaacaaa ttgataagca atgctttttt ataatgccaa
     5221 ctttgtacaa aaaagcaggc tttaaagg
//
