LOCUS       sequence-292672-        4433 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AND gate with IPTG and aTc as inputs and sfGFP as output.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4433)
  AUTHORS   Rosado A, Cordero T, Rodrigo G
  TITLE     Binary addition in a living cell based on riboregulation.
  JOURNAL   PLoS Genet. 2018 Jul 19;14(7):e1007548. doi: 
            10.1371/journal.pgen.1007548. eCollection 2018 Jul.
  PUBMED    30024870
REFERENCE   2  (bases 1 to 4433)
  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: "PLoS 
            Genet."; date: "2018-07-19"; volume: "14(7):e1007548. doi"; pages: "
            10.1371/journal.pgen.1007548. eCollection 2018 Jul"
COMMENT     Sequence Label: pRHA12
FEATURES             Location/Qualifiers
     source          1..4433
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    133..151
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    158..176
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    215..231
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             278..289
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             305..1015
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="RKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     misc_feature    1022..1042
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1078..1109
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(1208..2023)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1931..1950
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2015..2034
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(2381..3331)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFGIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      3379..3601
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(4093..4187)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      complement(4343..4372)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
ORIGIN
        1 ctagcatcga tgcgaaaaaa ccccgccgaa gcggggtttt ttgcggctag ccccagaatc
       61 ccttcgagca aatcctagcg aggtgccgct agattcgctc aaaggaaccc ttcttcctgc
      121 ccgtgctcag tatctctatc actgataggg atgtcaatct ctatcactga tagggaccta
      181 ggatgcatat aaatgtgagc ggataacatt gacattgtga gcggataaca agatactgag
      241 cacacccagt atcattctct tcttcctgcc cacgcggaaa gaggagaaag gtgtaatgac
      301 tagccgtaaa ggcgaagagc tgttcactgg tgtcgtccct attctggtgg aactggatgg
      361 tgatgtcaac ggtcataagt tttccgtgcg tggcgagggt gaaggtgacg caactaatgg
      421 taaactgacg ctgaagttca tctgtactac tggtaaactg ccggtacctt ggccgactct
      481 ggtaacgacg ctgacttatg gtgttcagtg ctttgctcgt tatccggacc atatgaagca
      541 gcatgacttc ttcaagtccg ccatgccgga aggctatgtg caggaacgca cgatttcctt
      601 taaggatgac ggcacgtaca aaacgcgtgc ggaagtgaaa tttgaaggcg ataccctggt
      661 aaaccgcatt gagctgaaag gcattgactt taaagaagac ggcaatatcc tgggccataa
      721 gctggaatac aattttaaca gccacaatgt ttacatcacc gccgataaac aaaaaaatgg
      781 cattaaagcg aattttaaaa ttcgccacaa cgtggaggat ggcagcgtgc agctggctga
      841 tcactaccag caaaacactc caatcggtga tggtcctgtt ctgctgccag acaatcacta
      901 tctgagcacg caaagcgttc tgtctaaaga tccgaacgag aaacgcgatc atatggttct
      961 gctggagttc gtaaccgcag cgggcatcac gcatggtatg gatgaactgt acaaatgatg
     1021 atactagtag cggccgctgc aggagtcact aagggttagt tagttagatt agcagaaagt
     1081 caaaagcctc cgaccggagg cttttgacta aaacttccct tggggttatc attggggctc
     1141 actcaaaggc ggtaatcaga taaaaaaaat ccttagcttt cgctaaggat gatttctgct
     1201 agtattatta gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat
     1261 tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc
     1321 agttccaaag aatggcaagg tcctggtaac ggtctgcgat tccgacccgt ccaacatcaa
     1381 tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag
     1441 tgacgactga atccggtgag aatggcaaga gcttgtgcat ttctttccag acttgttcaa
     1501 caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcatgc
     1561 gtgattgcgc ctgagcaaga cgaaatacac gatcgctgtt aaaaggacaa ttacaaacag
     1621 gaatcgaatg taaccggcgc aggaacacgg ccagcgcatc aacaatattt tcacctgaat
     1681 caggatattc ttctaatacc tggaaggctg ttttcccagg aatcgcggtg gtgagtaacc
     1741 acgcatcatc aggagtacgg ataaaatgct tgatggtcgg gagaggcata aactccgtca
     1801 gccagttgag acggaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt
     1861 tcagaaacaa ctctggcgca tcgggcttcc catacaagcg atagattgtc gcacctgatt
     1921 gcccgacatt atcgcgagcc catttatacc catataaatc agcgtccatg ttggagttta
     1981 agcgcggacg ggagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac
     2041 tgtttatgta agcagacagt tttattgttc atgatgatat atttttatct tgtgcaatgt
     2101 aacatcagag attttgagac acaacgtggc tttgttgaat aaatcgaact tttgctgagt
     2161 tgaaggatca gctctagtag ttacattgtc gatctgttca tggtgaacag ctttgaatgc
     2221 accaaaaact cgtaaaagct ctgatgtatc tatctttttt acaccgtttt catctgtgca
     2281 tatggacagt tttccctttg atatgtaacg gtgaacagtt gttctacttt tgtttgttag
     2341 tcttgatgct tcactgatag atacaagagc cataagaacc tcagatcctt ccgtatttag
     2401 ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag ccatgagaac gaaccattga
     2461 gatcatactt actttgcatg tcactcaaaa attttgcctc aaaactggtg agctgaattt
     2521 ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt atgtaggtag gaatctgatg
     2581 taatggttgt tggtattttg tcaccattca tttttatctg gttgttctca agttcggtta
     2641 cgagatccat ttgtctatct agttcaactt ggaaaatcaa cgtatcagtc gggcggcctc
     2701 gcttatcaac caccaatttc atattgctgt aagtgtttaa atctttactt attggtttca
     2761 aaacccattg gttaagcctt ttaaactcat ggtagttatt ttcaagcatt aacatgaact
     2821 taaattcatc aaggctaatc tctatatttg ccttgtgagt tttcttttgt gttagttctt
     2881 ttaataacca ctcataaatc ctcatagagt atttgttttc aaaagactta acatgttcca
     2941 gattatattt tatgaatttt tttaactgga aaagataagg caatatctct tcactaaaaa
     3001 ctaattctaa tttttcgctt gagaacttgg catagtttgt ccactggaaa atcccaaagc
     3061 ctttaaccaa aggattcctg atttccacag ttctcgtcat cagctctctg gttgctttag
     3121 ctaatacacc ataagcattt tccctactga tgttcatcat ctgagcgtat tggttataag
     3181 tgaacgatac cgtccgttct ttccttgtag ggttttcaat cgtggggttg agtagtgcca
     3241 cacagcataa aattagcttg gtttcatgct ccgttaagtc atagcgacta atcgctagtt
     3301 catttgcttt gaaaacaact aattcagaca tacatctcaa ttggtctagg tgattttaat
     3361 cactatacca attgagatgg gctagtcaat gataattaca tgtccttttc ctttgagttg
     3421 tgggtatctg taaattctgc tagacctttg ctggaaaact tgtaaattct gctagaccct
     3481 ctgtaaattc cgctagacct ttgtgtgttt tttttgttta tattcaagtg gttataattt
     3541 atagaataaa gaaagaataa aaaaagataa aaagaataga tcccagccct gtgtataact
     3601 cactacttta gtcagttccg cagtattaca aaaggatgtc gcaaacgctg tttgctcctc
     3661 tacaaaacag accttaaaac cctaaaggct taagtagcac cctcgcaagc tcgggcaaat
     3721 cgctgaatat tccttttgtc tccgaccatc aggcacctga gtcgctgtct ttttcgtgac
     3781 attcagttcg ctgcgctcac ggctctggca gtgaatgggg gtaaatggca ctacaggcgc
     3841 cttttatgga ttcatgcaag gaaactaccc ataatacaag aaaagcccgt cacgggcttc
     3901 tcagggcgtt ttatggcggg tctgctatgt ggtgctatct gactttttgc tgttcagcag
     3961 ttcctgccct ctgattttcc agtctgacca cttcggatta tcccgtgaca ggtcattcag
     4021 actggctaat gcacccagta aggcagcggt atcatcaaca ggcttacccg tcttactgtc
     4081 cctagtgctt ggattctcac caataaaaaa cgcccggcgg caaccgagcg ttctgaacaa
     4141 atccagatgg agttctgagg tcattactgg atctatcaac aggagtccaa gcgagctcgt
     4201 aaacttggtc tgacagctct agctccggca aaaaaacggg caaggtgtca ccaccctgcc
     4261 ctttttcttt aaaaccgaaa agattacttc gcgtttgcca cctgacgtct aagaaaagga
     4321 atattcagca atttgcccgt gccgaagaaa ggcccacccg tgaaggtgag ccagtgagtt
     4381 gattgctacg taattagtta gttagccctt agtgactcga attcgcggat ccg
//
