LOCUS       sequence-400017-        7672 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SR latch circuit on pLW555 backbone: PLux-S3, PTet-P1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7672)
  AUTHORS   Lebovich M, Zeng M, Andrews LB
  TITLE     Algorithmic Programming of Sequential Logic and Genetic Circuits for
            Recording Biochemical Concentration in a Probiotic Bacterium.
  JOURNAL   ACS Synth Biol. 2023 Aug 15. doi: 10.1021/acssynbio.3c00232.
  PUBMED    37581922
REFERENCE   2  (bases 1 to 7672)
  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: "ACS Synth 
            Biol. 2023 Aug 15. doi: 10.1021/acssynbio.3c00232."
COMMENT     Sequence Label: pML4
FEATURES             Location/Qualifiers
     source          1..7672
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(399..1214)
                     /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"
     CDS             complement(399..1214)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /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     1122..1141
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1206..1225
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     protein_bind    2482..2500
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2507..2525
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        3331..3365
                     /label=BBa_J23105
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.24"
     RBS             3374..3385
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             4241..4252
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             4370..5452
                     /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     complement(4389..4408)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             5471..6094
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /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(5544..5563)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(7053..7072)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     rep_origin      complement(join(7437..7672,1..310))
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(7517..7536)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgcacgaac cccccgttca gtccgaccgc tgcgccttat ccggtaacta tcgtcttgag
       61 tccaacccgg aaagacatgc aaaagcacca ctggcagcag ccactggtaa ttgatttaga
      121 ggagttagtc ttgaagtcat gcgccggtta aggctaaact gaaaggacaa gttttggtga
      181 ctgcgctcct ccaagccagt tacctcggtt caaagagttg gtagctcaga gaaccttcga
      241 aaaaccgccc tgcaaggcgg ttttttcgtt ttcagagcaa gagattacgc gcagaccaaa
      301 acgatctcaa gaagatcatc ttattaaggg gtctgacgct cagtggaacg aaaaatcaat
      361 ctaaagtata tatgagtaaa cttggtctga cagttacctt agaaaaactc atcgagcatc
      421 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
      481 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
      541 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
      601 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
      661 agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
      721 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
      781 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
      841 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
      901 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
      961 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     1021 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     1081 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     1141 ccatataaat cagcatccat gttggaattt aatcgcggcc tggagcaaga cgtttcccgt
     1201 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     1261 catgatgata tatttttatc ttgtgcaatg tacatcagag attttgagac acaaccaatt
     1321 attgaaggcc tccctaacgg ggggcctttt tttgtttctg gtctcccgct tggagcgacg
     1381 tacggtggaa tctgattcgt taccaattga catgatacga aacgtaccgt atcgttaagg
     1441 ttacgatagc ttcttaccgg acctgtagga tcgtacaggt ttacgcaaga aaatggtttg
     1501 ttactttcga ataaactgag cgctcaacgg gtgtgcttcc cgttctgatg agtccgtgag
     1561 gacgaaagcg cctctacaaa taattttgtt taagagtcta tggactatgt tttcacaaag
     1621 gaagtaccag gatggcacgt aaaaccgcag cagaagcaga agaaacccgt cagcgtatta
     1681 ttgatgcagc actggaagtt tttgttgcac agggtgttag tgatgcaacc ctggatcaga
     1741 ttgcacgtaa agccggtgtt acccgtggtg cagtttattg gcattttaat ggtaaactgg
     1801 aagttctgca ggcagttctg gcaagccgtc agcatccgct ggaactggat tttacaccgg
     1861 atctgggtat tgaacgtagc tgggaagcag ttgttgttgc aatgctggat gcagttcata
     1921 gtccgcagag caaacagttt agcgaaattc tgatttatca gggtctggat gaaagcggtc
     1981 tgattcataa tcgtatggtt caggcaagcg atcgttttct gcagtatatt catcaggttc
     2041 tgcgtcatgc agttacccag ggtgaactgc cgattaatct ggatctgcag accagcattg
     2101 gtgtttttaa aggtctgatt accggtctgc tgtatgaagg tctgcgtagc aaagatcagc
     2161 aggcacagat tatcaaagtt gcactgggta gcttttgggc actgctgcgt gaaccgcctc
     2221 gttttctgct gtgtgaagaa gcacagatta aacaggtgaa atccttcgaa taattcagcc
     2281 aaaaaactta agaccgccgg tcttgtccac taccttgcag taatgcggtg gacaggatcg
     2341 gcggttttct tttctcttct caaaggtcgc tggagtctat gattggtcca gattcgttac
     2401 caattgacag ctagctcagt cctaggtata tacatacatg cttgtttgtt tgtaaactac
     2461 gtactccacc gttggctttt ttccctatca gtgatagaga ttgacatccc tatcagtgat
     2521 agagataatg agcacctgag cggtcaacgc atgtgctttg cgttctgatg agacagtgat
     2581 gtcgaaaccg cctctacaaa taattttgtt taactatgga ctatgtttga aagggagaaa
     2641 tactagatgg cacgtacccc gagccgtagc agcattggta gcctgcgtag tccgcatacc
     2701 cataaagcaa ttctgaccag caccattgaa atcctgaaag aatgtggtta tagcggtctg
     2761 agcattgaaa gcgttgcacg tcgtgccggt gcaagcaaac cgaccattta tcgttggtgg
     2821 accaataaag cagcactgat tgccgaagtg tatgaaaatg aaagcgaaca ggtgcgtaaa
     2881 tttccggatc tgggtagctt taaagccgat ctggattttc tgctgcgtaa tctgtggaaa
     2941 gtttggcgtg aaaccatttg tggtgaagca tttcgttgtg ttattgcaga agcacagctg
     3001 gaccctgcaa ccctgaccca gctgaaagat cagtttatgg aacgtcgtcg tgagatgccg
     3061 aaaaaactgg ttgaaaatgc cattagcaat ggtgaactgc cgaaagatac caatcgtgaa
     3121 ctgctgctgg atatgatttt tggtttttgt tggtatcgcc tgctgaccga acagctgacc
     3181 gttgaacagg atattgaaga atttaccttc ctgctgatta atggtgtttg tccgggtaca
     3241 cagcgttaag gaaacacaga aaaaagcccg cacctgacag tgcgggcttt ttttttcgac
     3301 caaaggaggt aatggtcacc atatatcaag tttacggcta gctcagtcct aggtactatg
     3361 ctagctacta gagaaagagg agaaatacta gatggctgaa gcgcaaaatg atcccctgct
     3421 gccgggatac tcgtttaatg cccatctggt ggcgggttta acgccgattg aggccaacgg
     3481 ttatctcgat ttttttatcg accgaccgct gggaatgaaa ggttatattc tcaatctcac
     3541 cattcgcggt cagggggtgg tgaaaaatca gggacgagaa tttgtttgcc gaccgggtga
     3601 tattttgctg ttcccgccag gagagattca tcactacggt cgtcatccgg aggctcgcga
     3661 atggtatcac cagtgggttt actttcgtcc gcgcgcctac tggcatgaat ggcttaactg
     3721 gccgtcaata tttgccaata cggggttctt tcgcccggat gaagcgcacc agccgcattt
     3781 cagcgacctg tttgggcaaa tcattaacgc cgggcaaggg gaagggcgct attcggagct
     3841 gctggcgata aatctgcttg agcaattgtt actgcggcgc atggaagcga ttaacgagtc
     3901 gctccatcca ccgatggata atcgggtacg cgaggcttgt cagtacatca gcgatcacct
     3961 ggcagacagc aattttgata tcgccagcgt cgcacagcat gtttgcttgt cgccgtcgcg
     4021 tctgtcacat cttttccgcc agcagttagg gattagcgtc ttaagctggc gcgaggacca
     4081 acgtatcagc caggcgaagc tgcttttgag caccacccgg atgcctatcg ccaccgtcgg
     4141 tcgcaatgtt ggttttgacg atcaactcta tttctcgcgg gtatttaaaa aatgcaccgg
     4201 ggccagcccg agcgagttcc gtgccggtta ataaccaatt attgaaggcc gctaacgcag
     4261 cctttttttg tttctggtct cccaatggcg gcgcgccatc gaatggcgca aaacctttcg
     4321 cggtatggca tgatagcgcc cggaagagag tcaattcagg gtggtgaata tgaaaccagt
     4381 aacgttatac gatgtcgcag agtatgccgg tgtctcttat cagaccgttt cccgcgtggt
     4441 gaaccaggcc agccacgttt ctgcgaaaac gcgggaaaaa gtggaagcgg cgatggcgga
     4501 gctgaattac attcccaacc gcgtggcaca acaactggcg ggcaaacagt cgttgctgat
     4561 tggcgttgcc acctccagtc tggccctgca cgcgccgtcg caaattgtcg cggcgattaa
     4621 atctcgcgcc gatcaactgg gtgccagcgt ggtggtgtcg atggtagaac gaagcggcgt
     4681 cgaagcctgt aaagcggcgg tgcacaatct tctcgcgcaa cgcgtcagtg ggctgatcat
     4741 taactatccg ctggatgacc aggatgccat tgctgtggaa gctgcctgca ctaatgttcc
     4801 ggcgttattt cttgatgtct ctgaccagac acccatcaac agtattattt tctcccatga
     4861 ggacggtacg cgactgggcg tggagcatct ggtcgcattg ggtcaccagc aaatcgcgct
     4921 gttagcgggc ccattaagtt ctgtctcggc gcgtctgcgt ctggctggct ggcataaata
     4981 tctcactcgc aatcaaattc agccgatagc ggaacgggaa ggcgactgga gtgccatgtc
     5041 cggttttcaa caaaccatgc aaatgctgaa tgagggcatc gttcccactg cgatgctggt
     5101 tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt accgagtccg ggctgcgcgt
     5161 tggtgcggat atctcggtag tgggatacga cgataccgaa gatagctcat gttatatccc
     5221 gccgttaacc accatcaaac aggattttcg cctgctgggg caaaccagcg tggaccgctt
     5281 gctgcaactc tctcagggcc aggcggtgaa gggcaatcag ctgttgccag tctcactggt
     5341 gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     5401 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gacaatccag
     5461 gaggaaaaaa atgtccagat tagataaaag taaagtgatt aacagcgcat tagagctgct
     5521 taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga
     5581 gcagcctaca ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat
     5641 tgagatgtta gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga
     5701 ttttttacgt aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc
     5761 aaaagtacat ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt
     5821 agccttttta tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt
     5881 ggggcatttt actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga
     5941 aagggaaaca cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt
     6001 tgatcaccaa ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt
     6061 agaaaaacaa cttaaatgtg aaagtgggtc ctaaatcaca caggaaaggc ctcgatgaaa
     6121 aacataaatg ccgacgacac atacagaata attaataaaa ttaaagcttg tagaagcaat
     6181 aatgatatta atcaatgctt atctgatatg actaaaatgg tacattgtga atattattta
     6241 ctcgcgatca tttatcctca ttctatggtt aaatctgata tttcaatcct agataattac
     6301 cctaaaaaat ggaggcaata ttatgatgac gctaatttaa taaaatatga tcctatagta
     6361 gattattcta actccaatca ttcaccaatt aattggaata tatttgaaaa caatgctgta
     6421 aataaaaaat ctccaaatgt aattaaagaa gcgaaaacat caggtcttat cactgggttt
     6481 agtttcccta ttcatacggc taacaatggc ttcggaatgc ttagttttgc acattcagaa
     6541 aaagacaact atatagatag tttattttta catgcgtgta tgaacatacc attaattgtt
     6601 ccttctctag ttgataatta tcgaaaaata aatatagcaa ataataaatc aaacaacgat
     6661 ttaaccaaaa gagaaaaaga atgtttagcg tgggcatgcg aaggaaaaag ctcttgggat
     6721 atttcaaaaa tattaggttg cagtgagcgt actgtcactt tccatttaac caatgcgcaa
     6781 atgaaactca atacaacaaa ccgctgccaa agtatttcta aagcaatttt aacaggagca
     6841 attgattgcc catactttaa aaattaacca taattggtaa cgaatcagac aattgacggc
     6901 tcgagggagt agcatagggt ttgcagaatc cctgcttcgt ccatttgaca ggcacattat
     6961 gcatcgatga taagctgtca aacatgagca gatcctctac gccggacgca tcgtggccgg
     7021 catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga
     7081 agatcgggct cgccacttcg ggctcatgag caaatatttt atctgaggtg cttcctcgct
     7141 cactgactcg ctgcacgagg cagacctcag cgctagcgga gtgtatactg gcttactatg
     7201 ttggcactga tgagggtgtc agtgaagtgc ttcatgtggc aggagaaaaa aggctgcacc
     7261 ggtgcgtcag cagaatatgt gatacaggat atattccgct tcctcgctca ctgactcgct
     7321 acgctcggtc gttcgactgc ggcgagcgga aatggcttac gaacggggcg gagatttccc
     7381 ggaagatgcc aggaagatac ttaacaggga agtgagaggg ccgcggcaaa gccgtttttc
     7441 cataggctcc gcccccctga caagcatcac gaaatctgac gctcaaatca gtggtggcga
     7501 aacccgacag gactataaag ataccaggcg tttccccctg gcggctccct cgtgcgctct
     7561 cctgttcctg cctttcggtt taccggtgtc attccgctgt tatggccgcg tttgtctcat
     7621 tccacgcctg acactcagtt ccgggtaggc agttcgctcc aagctggact gt
//
