LOCUS       sequence-370628-        4935 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Molecular IMPLY gate, encodes mutant Ci protein from Plteto-1 
            promoter, and EGFP form PR and PlTataa promoter. Must be expressed 
            in E.coli DH5alphaz1 for experimental characterization..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4935)
  AUTHORS   Mukhopadhyay S, Sarkar K, Srivastava R, Pal A, Bagh S
  TITLE     Processing two environmental chemical signals with a synthetic 
            genetic IMPLY gate, a 2-input-2-output integrated logic circuit, and
            a process pipeline to optimize its systems chemistry in Escherichia 
            coli.
  JOURNAL   Biotechnol Bioeng. 2020 May;117(5):1502-1512. doi: 
            10.1002/bit.27286. Epub 2020 Feb 7.
  PUBMED    31981217
REFERENCE   2  (bases 1 to 4935)
  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"; doi: "10.1002/bit.27286";
            journalName: "Biotechnol Bioeng"; date: "2020-05"; volume: "117"; 
            issue: "5"; pages: "1502-1512"
COMMENT     Sequence Label: pNB_EA_R1CiM_4EGFP_R3EGFPi2
FEATURES             Location/Qualifiers
     source          1..4935
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        720..824
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(825..893,894..1685)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1043..1062)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      1715..1809
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      1897..2485
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2386..2405
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2758..2844)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(2856..3569,3570..3572,3573..3575))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(2859..3569,3570..3572,3573..3575))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     RBS             3582..3593
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     protein_bind    3605..3621
                     /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)."
     protein_bind    3629..3645
                     /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)."
     protein_bind    3651..3669
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3683..3699
                     /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)."
     protein_bind    3706..3724
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        join(3808..3826,3827..3832,3833..3849,3850..3855,
                     3856..3881)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    3808..3826
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3833..3851
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             3891..3902
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             3914..4621
                     /codon_start=1
                     /gene="cI"
                     /product="phage lambda repressor"
                     /label=lambda repressor
                     /translation="TKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQSG
                     VGALFNGINALNAYNAALLAKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEYPV
                     FSHVQAGMFSPELRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPDGM
                     LILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSVVG
                     KVIASQWPEETFG"
     terminator      complement(4636..4682)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(4706..4935,1..484,485..487,488..490))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(4709..4935,1..484,485..487,488..490))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
ORIGIN
        1 ccttgatgcc gttcttctgc ttgtcggcca tgatatagac gttgtggctg ttgtagttgt
       61 actccagctt gtgccccagg atgttgccgt cctccttgaa gtcgatgccc ttcagctcga
      121 tgcggttcac cagggtgtcg ccctcgaact tcacctcggc gcgggtcttg tagttgccgt
      181 cgtccttgaa gaagatggtg cgctcctgga cgtagccttc gggcatggcg gacttgaaga
      241 agtcgtgctg cttcatgtgg tcggggtagc ggctgaagca ctgcacgccg taggtcaggg
      301 tggtcacgag ggtgggccag ggcacgggca gcttgccggt ggtgcagatg aacttcaggg
      361 tcagcttgcc gtaggtggca tcgccctcgc cctcgccgga cacgctgaac ttgtggccgt
      421 ttacgtcgcc gtccagctcg accaggatgg gcaccacccc ggtgaacagc tcctcgccct
      481 tgctcaccat ggtacctttc tcgtctgctg tgaattcgta catgcaacca ttatcaccgc
      541 cagaggtaaa atagtcaaca cgcacggtgt tactcgaggt gaagacgaaa gggcctcgtg
      601 atacgcctat ttttataggt taatgtcatg ataataaaga tcttgatgtt ggcgtcttcg
      661 ttgtcgactg ttcgtcttcg tcccagtaga cgtcaggtgg cacttttcgg ggaaatgtgc
      721 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
      781 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
      841 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
      901 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
      961 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     1021 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     1081 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     1141 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     1201 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     1261 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     1321 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     1381 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     1441 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     1501 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     1561 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     1621 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     1681 ggtaactgtc agaccaagtt tacgagctcg cttggactcc tgttgataga tccagtaatg
     1741 acctcagaac tccatctgga tttgttcaga acgctcggtt gccgccgggc gttttttatt
     1801 ggtgagaatc caagcactag tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     1861 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     1921 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     1981 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     2041 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     2101 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     2161 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     2221 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     2281 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     2341 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     2401 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     2461 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     2521 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     2581 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacgg ttgttttgga
     2641 gcacggaact gcagaatgaa tagcttctgc caggatcctg actggcgacg taatccacga
     2701 aaatctatac attaagacga tctctttttc cgtcatcgcc taggtctagg gcggcggatt
     2761 tgtcctactc aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttt
     2821 cgactgagcc tttcgtttta tttgatgcct ctagattact tgtacagctc gtccatgccg
     2881 agagtgatcc cggcggcggt cacgaactcc agcaggacca tgtgatcgcg cttctcgttg
     2941 gggtctttgc tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg
     3001 ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga gctgcacgct gccgtcctcg
     3061 atgttgtggc ggatcttgaa gttcaccttg atgccgttct tctgcttgtc ggccatgata
     3121 tagacgttgt ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc
     3181 ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc
     3241 tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag
     3301 ccttcgggca tggcggactt gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg
     3361 aagcactgca cgccgtaggt cagggtggtc acgagggtgg gccagggcac gggcagcttg
     3421 ccggtggtgc agatgaactt cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg
     3481 ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag gatgggcacc
     3541 accccggtga acagctcctc gcccttgctc accatggtac ctttctcctc tttaatgaat
     3601 tcaattgtta tccgctcaca attatcaatt gttatccgct cacaattatc tctctatcac
     3661 tgatagggat gtgctcagta tcttgttatc cgctcacaat gtcaatctct atcactgata
     3721 gggactcgag gtgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     3781 tgataataaa gatcctggta actcgagtcc ctatcagtga tagagattga catccctatc
     3841 agtgatagag atactgagca catcagcagg acgcactgac cgaattcatt aaagaggaga
     3901 aaggtaccat ggcacaaaaa agaaaccatt aacacaagag cagcttgagg acgcacgtcg
     3961 ccttaaagca atttatgaaa aaaagaaaaa tgaacttggc ttatcccagg aatctgtcgc
     4021 agacaagatg gggatggggc agtcaggcgt tggtgcttta tttaatggca tcaatgcatt
     4081 aaatgcttat aacgccgcat tgcttgcaaa aattctcaaa gttagcgttg aagaatttag
     4141 cccttcaatc gccagagaaa tctacgagat gtatgaagcg gttagtatgc agccgtcact
     4201 tagaagtgag tatgagtacc ctgttttttc tcatgttcag gcagggatgt tctcacctga
     4261 gcttagaacc tttaccaaag gtgatgcgga gagatgggta agcacaacca aaaaagccag
     4321 tgattctgca ttctggcttg aggttgaagg taattccatg accgcaccaa caggctccaa
     4381 gccaagcttt cctgacggaa tgttaattct cgttgaccct gagcaggctg ttgagccagg
     4441 tgatttctgc atagccagac ttgggggtga tgagtttacc ttcaagaaac tgatcaggga
     4501 tagcggtcag gtgtttttac aaccactaaa cccacagtac ccaatgatcc catgcaatga
     4561 gagttgttcc gttgtgggga aagttatcgc tagtcagtgg cctgaagaga cgtttggctg
     4621 atctagaggc atcaaataaa acgaaaggcc cagtctttcg actgagcctt tcgttttatt
     4681 tgatgcctct agagtcgcgg ccgctttact tgtacagctc gtccatgccg agagtgatcc
     4741 cggcggcggt cacgaactcc agcaggacca tgtgatcgcg cttctcgttg gggtctttgc
     4801 tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga
     4861 tgggggtgtt ctgctggtag tggtcggcga gctgcacgct gccgtcctcg atgttgtggc
     4921 ggatcttgaa gttca
//
