LOCUS       sequence-297010-        5795 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Toggle switch circuit responsive to AHL gradient..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5795)
  AUTHORS   Barbier I, Perez-Carrasco R, Schaerli Y
  TITLE     Controlling spatiotemporal pattern formation in a concentration 
            gradient with a synthetic toggle switch.
  JOURNAL   Mol Syst Biol. 2020 Jun;16(6):e9361. doi: 10.15252/msb.20199361.
  PUBMED    32529808
REFERENCE   2  (bases 1 to 5795)
  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: "Mol Syst 
            Biol."; date: "2020-06"; volume: "16(6):e9361. doi"; pages: " 
            10.15252/msb.20199361"
COMMENT     Sequence Label: TS_pLuxLac
FEATURES             Location/Qualifiers
     source          1..5795
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(63..82)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(658..752)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(783..1577)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(894..913)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1504..1523
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(1771..2484)
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1811..1831)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     2426..2454
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     promoter        complement(join(2536..2561,2562..2567,2568..2584,2585..2590,
                     2591..2609))
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    2566..2584
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2591..2609
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             complement(2652..3734)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     3696..3715
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(join(3771..3796,3797..3802,3803..3819,3820..3825,
                     3826..3844))
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    3801..3819
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3826..3844
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3887..3903
                     /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    3918..3934
                     /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)."
     CDS             3976..4599
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(4049..4068)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    4666..4682
                     /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    4697..4713
                     /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)."
     CDS             4759..5469
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4904..4922)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5172..5191
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     terminator      5529..5614
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(join(5778..5795,1..571))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
       61 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
      121 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
      181 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtctgca
      241 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
      301 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
      361 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
      421 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
      481 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
      541 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
      601 tgacgctcag tggaacgaaa actcacgtta agggatttgg tcatggctag tgcttggatt
      661 ctcaccaata aaaaacgccc ggcggcaacc gagcgttctg aacaaatcca gatggagttc
      721 tgaggtcatt actggatcta tcaacaggag tccaagccaa ttctcgaacc ccagagtccc
      781 gctcagaaga actcgtcaag aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg
      841 ataccgtaaa gcacgaggaa gcggtcagcc cattcgccgc caagctcttc agcaatatca
      901 cgggtagcca acgctatgtc ctgatagcgg tccgccacac ccagccggcc acagtcgatg
      961 aatccagaaa agcggccatt ttccaccatg atattcggca agcaggcatc gccgtgggtc
     1021 acgacgagat cctcgccgtc gggcatacgc gccttgagcc tggcgaacag ttcggctggc
     1081 gcgagcccct gatgctcttc gtccagatca tcctgatcga caagaccggc ttccatccga
     1141 gtacgtgctc gctcgatgcg atgtttcgct tggtggtcga atgggcaggt agccggatca
     1201 agcgtatgca gccgccgcat tgcatcagcc atgatggata ctttctcggc aggagcaagg
     1261 tgagatgaca ggagatcctg ccccggcact tcgcccaata gcagccagtc ccttcccgct
     1321 tcagtgacaa cgtcgagcac agccgcgcaa ggaacgcccg tcgtggccag ccacgatagc
     1381 cgcgctgcct cgtcttgcag ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga
     1441 accgggcgac cctgcgctga cagccggaac acggcggcat cagagcagcc gattgtctgt
     1501 tgtgcccagt catagccgaa tagcctctcc acccaagccg ccggagaacc tgcgtgcaat
     1561 ccatcttgtt caatcatgcg aaacgatcct catcctgtct cttgatcaga tcttgatccc
     1621 ctgcgccatc agatccttgg cggcaagaaa gccatccagt ttactttgca gggcttccca
     1681 accttaccag agggcggccc aactggcaat tccgacgtct gtgcaagtac tactgttctg
     1741 cagtcacttg aattcgatac ccagctgtta tttgtatagt tcatccatgc catgtgtaat
     1801 cccagcagcg gttacaaact caagaaggac catgtggtct ctcttttcgt tgggatcttt
     1861 cgaaagggca gattgtgtgg acaggtaatg gttgtctggt aaaaggacag ggccatcgcc
     1921 aatcggagta ttttgttgat aatggtctgc tagttgaacg cttccatctt caatgttgtg
     1981 tctaattttg aagttaactt tgattccatt cttttgtttg tctgccatga tgtatacatt
     2041 gtgtgagtta tagttgtatt ccaatttgtg tccaagaatg tttccatctt ctttaaaatc
     2101 aatacctttt aactcgattc tattaacaag ggtatcacct tcaaacttga cttcagcacg
     2161 tgtcttgtag ttcccgtcat ctttgaaaaa tatagttctt tcctgtacgt aaccttcggg
     2221 catggcactc ttgaaaaagt catgctgttt catgtgatct gggtatctcg caaagcattg
     2281 aacaccataa ccgaaagtag tgacaagtgt tggccacgga acaggtagtt ttccagtagt
     2341 gcaaataaat ttaagggtaa gttttccgta tgttgcatca ccttcaccct ctccactgac
     2401 agaaaatttg tgcccattaa catcaccatc taattcaaca agaattggga caactccagt
     2461 gaaaagttct tctcctttac gcatgtgcac tacctccttg gttgtctatg ctatgctgat
     2521 ctacaactgg catgcggtca gtgcgtcctg ctgatgtgct cagtatctct atcactgata
     2581 gggatgtcaa tctctatcac tgatagggaa acgtttcgca gaagcttccg caaggtacca
     2641 ctttgccgcg gtcactgccc gctttccagt cgggaaacct gtcgtgccag ttgcattaat
     2701 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gagccagggt ggtttttctt
     2761 ttcaccagtg agacgggcaa cagttgattg cccttcaccg cctggccctg agagagttgc
     2821 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     2881 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatgtccgca
     2941 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     3001 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     3061 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     3121 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     3181 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     3241 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     3301 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     3361 ttaatgatca gcccactgac gcgctgcgcg agaagattgt gaaccgccgc tttacaggct
     3421 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     3481 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     3541 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     3601 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     3661 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtacagc
     3721 gttactggtt tcattgtaca ccctctcctt atattgggac ctatggatcc ggtcagtgcg
     3781 tcctgctgat gtgctcagta tctctatcac tgatagggat gtcaatctct atcactgata
     3841 gggacggccg cccgttccat ggacctgtag gatcgtacag gtttacttgt gagcggataa
     3901 caatatagtg tgtggaattg tgagcggata acaattagtc gaccgccttc ggcgaagcta
     3961 gggacgagag ctagcatgtc tcgtttagat aaaagtaaag tgattaacag cgcattagag
     4021 ctgcttaatg aggtcggaat cgaaggttta acaacccgta aactcgccca gaagctaggt
     4081 gtagagcagc ctacattgta ttggcatgta aaaaataagc gggctttgct cgacgcctta
     4141 gccattgaga tgttagatag gcaccatact cacttttgcc ctttagaagg ggaaagctgg
     4201 caagattttt tacgtaataa cgctaaaagt tttagatgtg ctttactaag tcatcgcgat
     4261 ggagcaaaag tacatttagg tacacggcct acagaaaaac agtatgaaac tctcgaaaat
     4321 caattagcct ttttatgcca acaaggtttt tcactagaga atgcattata tgcactcagc
     4381 gctgtggggc attttacttt aggttgcgta ttggaagatc aagagcatca agtcgctaaa
     4441 gaagaaaggg aaacacctac tactgatagt atgccgccat tattacgaca agctatcgaa
     4501 ttatttgatc accaaggtgc agagccagcc ttcttattcg gccttgaatt gatcatctgc
     4561 ggattagaaa aacaacttaa atgtgaaagt gggtcttgag agctcggact gcttaagtcg
     4621 ctccatatgc tcgttcccgg gacctgtagg atcgtacagg tttacttgtg agcggataac
     4681 aatatagtgt gtggaattgt gagcggataa caattacaat tgagtattaa ctatcgttca
     4741 actgataggg aggcgccgat ggtgagcaag ggcgaggagg ataacatggc catcatcaag
     4801 gagttcatgc gcttcaaggt tcacatggag ggctccgtga acggccacga gttcgagatc
     4861 gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc
     4921 aagggtggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc
     4981 aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag
     5041 ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag
     5101 gactcctccc tgcaagacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc
     5161 ccctccgacg gccccgtaat gcagaagaag actatgggct gggaggcctc ctccgagcgg
     5221 atgtaccccg aggacggcgc gctgaagggc gagatcaagc agaggctgaa gctgaaggac
     5281 ggcggccact acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcaactg
     5341 cccggcgcgt acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc
     5401 atcgtggaac agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg
     5461 tataagtaag atatctatcg ccctagggac cgtctcgaga gaatcaatat taatccaacg
     5521 cgtggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
     5581 gtttgtcggt gaacgctctc ctgagtagga caaaccgccg ccctagactt aggcgttcgg
     5641 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     5701 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     5761 gccgcgttgc tggcgttttt ccataggctc cgccc
//
