LOCUS       sequence-201648-        5532 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses thsR from a TetR regulated promoter under ATC control. 
            Expresses sfGFP from a thsR regulated promoter. Expresses mCherry 
            from a constituative promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5532)
  AUTHORS   Landry BP, Palanki R, Dyulgyarov N, Hartsough LA, Tabor JJ
  TITLE     Phosphatase activity tunes two-component system sensor detection 
            threshold.
  JOURNAL   Nat Commun. 2018 Apr 12;9(1):1433. doi: 10.1038/s41467-018-03929-y.
  PUBMED    29650958
REFERENCE   2  (bases 1 to 5532)
  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: "Nat 
            Commun."; date: "2018-04-12"; volume: "9(1):1433. doi"; pages: " 
            10.1038/s41467-018-03929-y"
COMMENT     Sequence Label: pBL119-2
FEATURES             Location/Qualifiers
     source          1..5532
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(152..740)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(232..251)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(828..922)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(943..1602)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1517..1536
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        1731..1765
                     /label=BBa_J23114
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.1"
     RBS             1775..1786
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1794..2504
                     /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(1939..1957)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2207..2226
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2605..2623
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(2644..2662,2663..2668,2669..2685,2686..2691,
                     2692..2717)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    2644..2662
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2669..2687
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      3378..3449
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3465..3492
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             3808..3819
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             3826..4539
                     /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
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      4566..4637
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4653..4680
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        4732..4766
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
     CDS             4801..5427
                     /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(4877..4896)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      5433..5519
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 cctagggcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
       61 ccacagaatc aggggataac gcatgcaaga acatgtgagc aaaaggccag caaaaggcca
      121 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
      181 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
      241 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
      301 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
      361 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
      421 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
      481 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
      541 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
      601 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
      661 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
      721 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
      781 ggaacgaaaa ctcacgttaa gggattttgg tcatgactag tgcttggatt ctcaccaata
      841 aaaaacgccc ggcggcaacc gagcgttctg aacaaatcca gatggagttc tgaggtcatt
      901 actggatcta tcaacaggag tccaagcgag ctcgatatca aattacgccc cgccctgcca
      961 ctcatcgcag tactgttgta attcattaag cattctgccg acatggaagc catcacaaac
     1021 ggcatgatga acctgaatcg ccagcggcat cagcaccttg tcgccttgcg tataatattt
     1081 gcccatggtg aaaacggggg cgaagaagtt gtccatattg gccacgttta aatcaaaact
     1141 ggtgaaactc acccagggat tggctgagac gaaaaacata ttctcaataa accctttagg
     1201 gaaataggcc aggttttcac cataacacgc cacatcttgc gaatatatgt gtagaaactg
     1261 ccggaaatcg tcgtggtatt cactccagag cgatgaaaac gtttcagttt gctcatggaa
     1321 aacggtgtaa caagggtgaa cactatccca tatcaccagc tcaccgtctt tcattgccat
     1381 acgaaattcc ggatgagcat tcatcaggcg ggcaagaatg tgaataaagg ccggataaaa
     1441 cttgtgctta tttttcttta cggtctttaa aaaggccgta atatccagct gaacggtctg
     1501 gttataggta cattgagcaa ctgactgaaa tgcctcaaaa tgttctttac gatgccattg
     1561 ggatatatca acggtggtat atccagtgat ttttttctcc attttagctt ccttagctcc
     1621 tgaaaatctc gataactcaa aaaatacgac cggtagtgat cttatttcat tatggtgaaa
     1681 gttggaacct cttacgggcc ggccaacgtc tcattttcgc cagatatcga tttatggcta
     1741 gctcagtcct aggtacaatg ctagcgaatc cattaaagag gagaaaggta cccatggtga
     1801 gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca
     1861 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
     1921 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct
     1981 gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg
     2041 acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga
     2101 acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt
     2161 tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga
     2221 agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac ggcgccctga
     2281 agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca
     2341 agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcaacatca
     2401 agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg
     2461 agggccgcca ctccaccggc ggcatggacg agctgtacaa gtaagctttc taactaaaaa
     2521 ggcctcccaa atcggggggc cttttttatt gataacaaaa cgctcgtcta agaaaccatt
     2581 attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg tcttcacctc
     2641 gagtccctat cagtgataga gattgacatc cctatcagtg atagagatac tgagcacatc
     2701 agcaggacgc actgaccttc catatagcgt aatctaaaag attacattat gcagcagcaa
     2761 atcaacggcc cggtctacct ggtggatgat gatgaagcca ttatcgactc catcgatttt
     2821 ttgatggagg gctacggtta caaactgaac tcgtttaact gcggcgatcg ctttttggca
     2881 gaagtcgatc tcacccaggc aggatgtgta attctggatg cgcgtatgcc aggcttaact
     2941 ggtcctcagg tgcaacagct gctgagcgac gcgaaaagcc cgcttgcggt catcttcctg
     3001 accggccatg gcgatgttcc gatggcggtt gatgcgttca aaaatggcgc gttcgatttc
     3061 tttcaaaaac ctgtgccggg tagcttgctc agtcagtcaa ttgccaaagg cttgacttat
     3121 tcaatcgatc aacatctgaa acgtactaac caagcgttaa tcgacacgct ctcggaacgc
     3181 gaagctcaaa tttttcaact ggtgattgca ggcaacacca acaaacagat ggctaacgag
     3241 ctttgcgtgg ctattcgtac cattgaggtt caccgtagca aactgatgac caaactgggt
     3301 gttaacaacc tggctgaact ggttaaactg gcgccgctgc tggcacataa atccgaataa
     3361 taatctagac caggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt
     3421 ttatctgttg tttgtcggtg aacgctctct actagagtca cactggctca ccttcgggtg
     3481 ggccttcctg cgtttatagt aaatcactgc ataattcgtg tcgctcaagg cgcactcccg
     3541 ttctggataa tgttttttgc gccgacatca taacggttct ggcaaatatt ctgaaatgag
     3601 ctggttagct agtcagcaaa tttgaactgt gaattgcttc acaaacgccg ctatcgcaat
     3661 gtcagtatgt ggtttaccac aatatctaat atcactctgc tcaataacaa tgatgaaaac
     3721 cttaggaaga agttaattgt gttaaacagt taactagggg ctttatctaa cgctctccta
     3781 aggacaactg tcattgggag atttaacaaa gaggagaaat actagatgcg taaaggcgaa
     3841 gagctgttca ctggtgtcgt ccctattctg gtggaactgg atggtgatgt caacggtcat
     3901 aagttttccg tgcgtggcga gggtgaaggt gacgcaacta atggtaaact gacgctgaag
     3961 ttcatctgta ctactggtaa actgccggta ccttggccga ctctggtaac gacgctgact
     4021 tatggtgttc agtgttttgc tcgttatccg gaccatatga agcagcatga cttcttcaag
     4081 tccgccatgc cggaaggcta tgtgcaggaa cgcacgattt cctttaagga tgacggcacg
     4141 tacaaaacgc gtgcggaagt gaaatttgaa ggcgataccc tggtaaaccg cattgagctg
     4201 aaaggcattg actttaaaga agacggcaat atcctgggcc ataagctgga atacaatttt
     4261 aacagccaca atgtttacat caccgccgat aaacaaaaaa atggcattaa agcgaatttt
     4321 aaaattcgcc acaacgtgga ggatggcagc gtgcagctgg ctgatcacta ccagcaaaac
     4381 actccaatcg gtgatggtcc tgttctgctg ccagacaatc actatctgag cacgcaaagc
     4441 gttctgtcta aagatccgaa cgagaaacgc gatcatatgg ttctgctgga gttcgtaacc
     4501 gcagcgggca tcacgcatgg tatggatgaa ctgtacaaat gatgataata atctagacca
     4561 ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     4621 tgtcggtgaa cgctctctac tagagtcaca ctggctcacc ttcgggtggg cctttctgcg
     4681 tttataggta cccgtggatc ctctagagtc gacctgcagg catgcaagct ttttacggct
     4741 agctcagtcc taggtatagt gctagcccag ccagagaaac actctttaac agggggtagt
     4801 atgatgtctc gtttagataa aagtaaagtg attaacagcg cattagagct gcttaatgag
     4861 gtcggaatcg aaggtttaac aacccgtaaa ctcgcccaga agctaggtgt agagcagcct
     4921 acattgtatt ggcatgtaaa aaataagcgg gctttgctcg acgccttagc cattgagatg
     4981 ttagataggc accatactca cttttgccct ttagaagggg aaagctggca agatttttta
     5041 cgtaataacg ctaaaagttt tagatgtgct ttactaagtc atcgcgatgg agcaaaagta
     5101 catttaggta cacggcctac agaaaaacag tatgaaactc tcgaaaatca attagccttt
     5161 ttatgccaac aaggtttttc actagagaat gcattatatg cactcagcgc agtggggcat
     5221 tttactttag gttgcgtatt ggaagatcaa gagcatcaag tcgctaaaga agaaagggaa
     5281 acacctacta ctgatagtat gccgccatta ttacgacaag ctatcgaatt atttgatcac
     5341 caaggtgcag agccagcctt cttattcggc cttgaattga tcatatgcgg attagaaaaa
     5401 caacttaaat gtgaaagtgg gtcttaaggc atcaaataaa acgaaaggct cagtcgaaag
     5461 actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc
     5521 cgccgcccta ga
//
