LOCUS       sequence-178326-        5438 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses S. baltica TtrR under the PltetO-1 promoter, sfGFP under 
            PttrB185-269, and mCherry constitutively.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5438)
  AUTHORS   Daeffler KN, Galley JD, Sheth RU, Ortiz-Velez LC, Bibb CO, Shroyer 
            NF, Britton RA, Tabor JJ
  TITLE     Engineering bacterial thiosulfate and tetrathionate sensors for 
            detecting gut inflammation.
  JOURNAL   Mol Syst Biol. 2017 Apr 3;13(4):923. doi: 10.15252/msb.20167416.
  PUBMED    28373240
REFERENCE   2  (bases 1 to 5438)
  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: "2017-04-3"; volume: "13(4):923. doi"; pages: " 
            10.15252/msb.20167416"
COMMENT     Sequence Label: pKD233.7-3
FEATURES             Location/Qualifiers
     source          1..5438
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      1..87
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(252..840)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(332..351)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(928..1022)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1043..1702)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1617..1636
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        1831..1865
                     /label=BBa_J23114
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.1"
     RBS             1875..1886
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1894..2604
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPVDIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPSAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2039..2057)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2307..2326
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2705..2723
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(2744..2762,2763..2768,2769..2785,2786..2791,
                     2792..2817)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    2744..2762
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2769..2787
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      3491..3562
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3578..3605
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             3814..3825
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             3832..4545
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHMLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      4572..4643
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4659..4686
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        4738..4772
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
     CDS             4807..5433
                     /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="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(4883..4902)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
ORIGIN
        1 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg
       61 gtgaacgctc tcctgagtag gacaaatccg ccgccctaga cctagggcgt tcggctgcgg
      121 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
      181 gcatgcaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
      241 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
      301 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
      361 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
      421 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
      481 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
      541 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
      601 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
      661 aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg
      721 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
      781 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
      841 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
      901 gggattttgg tcatgactag tgcttggatt ctcaccaata aaaaacgccc ggcggcaacc
      961 gagcgttctg aacaaatcca gatggagttc tgaggtcatt actggatcta tcaacaggag
     1021 tccaagcgag ctcgatatca aattacgccc cgccctgcca ctcatcgcag tactgttgta
     1081 attcattaag cattctgccg acatggaagc catcacaaac ggcatgatga acctgaatcg
     1141 ccagcggcat cagcaccttg tcgccttgcg tataatattt gcccatggtg aaaacggggg
     1201 cgaagaagtt gtccatattg gccacgttta aatcaaaact ggtgaaactc acccagggat
     1261 tggctgagac gaaaaacata ttctcaataa accctttagg gaaataggcc aggttttcac
     1321 cataacacgc cacatcttgc gaatatatgt gtagaaactg ccggaaatcg tcgtggtatt
     1381 cactccagag cgatgaaaac gtttcagttt gctcatggaa aacggtgtaa caagggtgaa
     1441 cactatccca tatcaccagc tcaccgtctt tcattgccat acgaaattcc ggatgagcat
     1501 tcatcaggcg ggcaagaatg tgaataaagg ccggataaaa cttgtgctta tttttcttta
     1561 cggtctttaa aaaggccgta atatccagct gaacggtctg gttataggta cattgagcaa
     1621 ctgactgaaa tgcctcaaaa tgttctttac gatgccattg ggatatatca acggtggtat
     1681 atccagtgat ttttttctcc attttagctt ccttagctcc tgaaaatctc gataactcaa
     1741 aaaatacgac cggtagtgat cttatttcat tatggtgaaa gttggaacct cttacgggcc
     1801 ggccaacgtc tcattttcgc cagatatcga tttatggcta gctcagtcct aggtacaatg
     1861 ctagcgaatc cattaaagag gagaaaggta cccatggtga gcaagggcga ggaggataac
     1921 atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc
     1981 cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc
     2041 aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag
     2101 ttcatgtacg gctccaaggc ctacgtgaag caccccgtcg acatccccga ctacttgaag
     2161 ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg
     2221 gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg
     2281 cgcggcacca acttcccctc cgacggcccc gtaatgcaga agaagaccat gggctgggag
     2341 gcctcctccg agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg
     2401 ctgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag
     2461 aagcccgtgc agctgcccag cgcctacaac gtcaacatca agttggacat cacctcccac
     2521 aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc
     2581 ggcatggacg agctgtacaa gtaagctttc taactaaaaa ggcctcccaa atcggggggc
     2641 cttttttatt gataacaaaa cgctcgtcta agaaaccatt attatcatga cattaaccta
     2701 taaaaatagg cgtatcacga ggccctttcg tcttcacctc gagtccctat cagtgataga
     2761 gattgacatc cctatcagtg atagagatac tgagcacatc agcaggacgc actgaccact
     2821 ttaattcatt accacactta ataggagcga taatgagcct ggcgctgccg gtgtacctga
     2881 ttgacgacga tgactccgtt cgccgttccc tgcgtttcat gctggaaagc tacggcctga
     2941 aaatcactga cttcgattct gctgaagcgt tcttcaccgc ggtagacctg accctgccgg
     3001 gttgcgcact ggtggacgta cgtatgccgg gcctgagcgg cccacagctg cacctggaac
     3061 tggtgtctaa gaacagcccg ctggccgtga tctatctgac cggtcacggc gacgttccga
     3121 tggcggttga agcgctgaaa ctgggtgcgg tagatttctt tcagaaacct gcagacggcg
     3181 ctaaactggc tgaagctgtg gtcaaagccc tggaacacgc gaaaacccac taccaagaca
     3241 accagtacct ggaaacctat caggctctga ccccacgtga gcgcgaaatc ctgaacctga
     3301 ttgcgcaagg tctgaaaaac caggaaatcg cggacaacct gtgcattgcg atgcgcaccg
     3361 tagaaatcca ccgtgcgaac ctgatgaaag gtatgcaggt gggcagcctg gcagagatga
     3421 tgctgattta ctctcgtatc gcggagcgtc tgcgcctgga agataaccgt taataatcta
     3481 gaccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     3541 ttgtttgtcg gtgaacgctc tctactagag tcacactggc tcaccttcgg gtgggccttt
     3601 ctgcgtttat agtaaatcac tgcataattc gtgtcgctca aggcgcactc ccgttctgga
     3661 taatgttttt tgcgccgaca tcataacggt tctggcaaat attctgaaat gagctggtta
     3721 gctagtcaat atttgttgcg ctagatcaaa tccacgcctg atatgtggaa aaccactata
     3781 gttatgccgc ctcgcctttt acagaatgcc tgtaaagagg agaaatacta gatgcgtaaa
     3841 ggcgaagagc tgttcactgg tgtcgtccct attctggtgg aactggatgg tgatgtcaac
     3901 ggtcataagt tttccgtgcg tggcgagggt gaaggtgacg caactaatgg taaactgacg
     3961 ctgaagttca tctgtactac tggtaaactg ccggtacctt ggccgactct ggtaacgacg
     4021 ctgacttatg gtgttcagtg ctttgctcgt tatccggacc atatgaagca gcatgacttc
     4081 ttcaagtccg ccatgccgga aggctatgtg caggaacgca cgatttcctt taaggatgac
     4141 ggcacgtaca aaacgcgtgc ggaagtgaaa tttgaaggcg ataccctggt aaaccgcatt
     4201 gagctgaaag gcattgactt taaagaagac ggcaatatcc tgggccatat gctggaatac
     4261 aattttaaca gccacaatgt ttacatcacc gccgataaac aaaaaaatgg cattaaagcg
     4321 aattttaaaa ttcgccacaa cgtggaggat ggcagcgtgc agctggctga tcactaccag
     4381 caaaacactc caatcggtga tggtcctgtt ctgctgccag acaatcacta tctgagcacg
     4441 caaagcgttc tgtctaaaga tccgaacgag aaacgcgatc atatggttct gctggagttc
     4501 gtaaccgcag cgggcatcac gcatggtatg gatgaactgt acaaatgatg ataataatct
     4561 agaccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct
     4621 gttgtttgtc ggtgaacgct ctctactaga gtcacactgg ctcaccttcg ggtgggcctt
     4681 tctgcgttta taggtacccg tggatcctct agagtcgacc tgcaggcatg caagcttttt
     4741 acggctagct cagtcctagg tatagtgcta gcccagccag agaaacactc tttaacaggg
     4801 ggtagtatga tgtctcgttt agataaaagt aaagtgatta acagcgcatt agagctgctt
     4861 aatgaggtcg gaatcgaagg tttaacaacc cgtaaactcg cccagaagct aggtgtagag
     4921 cagcctacat tgtattggca tgtaaaaaat aagcgggctt tgctcgacgc cttagccatt
     4981 gagatgttag ataggcacca tactcacttt tgccctttag aaggggaaag ctggcaagat
     5041 tttttacgta ataacgctaa aagttttaga tgtgctttac taagtcatcg cgatggagca
     5101 aaagtacatt taggtacacg gcctacagaa aaacagtatg aaactctcga aaatcaatta
     5161 gcctttttat gccaacaagg tttttcacta gagaatgcat tatatgcact cagcgcagtg
     5221 gggcatttta ctttaggttg cgtattggaa gatcaagagc atcaagtcgc taaagaagaa
     5281 agggaaacac ctactactga tagtatgccg ccattattac gacaagctat cgaattattt
     5341 gatcaccaag gtgcagagcc agccttctta ttcggccttg aattgatcat atgcggatta
     5401 gaaaaacaac ttaaatgtga aagtgggtct taaggcat
//
