LOCUS       sequence-247897-        4802 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of torR(1-134aa)-ccaR(127-234aa), ompR137 chimera under 
            PLtetO-1, output Promoter PcpcG2-172.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4802)
  AUTHORS   Schmidl SR, Ekness F, Sofjan K, Daeffler KN, Brink KR, Landry BP, 
            Gerhardt KP, Dyulgyarov N, Sheth RU, Tabor JJ
  TITLE     Rewiring bacterial two-component systems by modular DNA-binding 
            domain swapping.
  JOURNAL   Nat Chem Biol. 2019 Jul;15(7):690-698. doi: 
            10.1038/s41589-019-0286-6. Epub 2019 May 20.
  PUBMED    31110305
REFERENCE   2  (bases 1 to 4802)
  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: 
            "10.1038/s41589-019-0286-6"; journalName: "Nat Chem Biol"; date: 
            "2019-07"; volume: "15"; issue: "7"; pages: "690-698"
COMMENT     Sequence Label: pKD263
FEATURES             Location/Qualifiers
     source          1..4802
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     30..48
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(69..87,88..93,94..110,111..116,117..142)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    69..87
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    94..112
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      927..998
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1014..1041
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             1337..1348
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1355..2068
                     /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      2095..2166
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2182..2209
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        2261..2295
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
     CDS             2330..2956
                     /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(2406..2425)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      2962..3048
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(3213..3801)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3293..3312)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(3889..3983)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(4004..4663)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4578..4597
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtcttc
       61 acctcgagtc cctatcagtg atagagattg acatccctat cagtgataga gatactgagc
      121 acatcagcag gacgcactga cctaagcgta aaataagtaa attacaccta tacaagtatg
      181 ccacatcaca ttgttattgt tgaagatgag ccggttaccc aggcgcgatt acaatcctac
      241 ttcactcagg aggggtatac cgtttccgtt acggcgagcg gtgctgggct acgggaaatt
      301 atgcagaatc agtcggtaga tttaattctg ctggatatca acttacccga tgaaaatggc
      361 ctgatgttaa cccgcgccct gcgagaacgc tcaacggtgg ggattattct ggttaccgga
      421 cgcagcgatc ggattgaccg tattgttggg ctggaaatgg gcgcagacga ttacgtcacc
      481 aaaccgctgg aactgcgcga actggtagta cgggtgaaaa atctgctctg gcgaatcgac
      541 ctcgcgcgac aagctcaacc gcacactcag gacaactgct tagagtgggg gccaatcagg
      601 ttggatccaa gcacctatga agttagttat gacaatgagg ttttgtcttt gacccgcaag
      661 gaatacagca ttctggaatt actactccgc aatggccgtc gggtgctaag tcggagcatg
      721 attatcgata gtatctggaa gttggagagt cccccagagg aagatacggt taaggtgcat
      781 gtgcggagtt tgcgacaaaa attaaaaagt gccggtttat cagcagatgc cattgaaacg
      841 gtccatggca ttgggtatcg tctggccaat ttaacggaaa aatctttgtg ccaagggaaa
      901 aactagtaat aatctagacc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg
      961 cctttcgttt tatctgttgt ttgtcggtga acgctctcta ctagagtcac actggctcac
     1021 cttcgggtgg gcctttctgc gtttatagta aatcactgca taattcgtgt cgctcaaggc
     1081 gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc
     1141 tgaaatgagc tggttagcta gtcaagccca ttgtgctttt ctctatcaac ctcagcttac
     1201 ctgaaggggt gaacaggtct gggttaattc atgttgcgaa atgtaacagt tttagtcgca
     1261 tcagctaact ttccgatttc tttacgattt tctccccctt ttcttcaatt ttactttgtt
     1321 aggatcgcat ttttaaaaag aggagaaata ctagatgcgt aaaggcgaag agctgttcac
     1381 tggtgtcgtc cctattctgg tggaactgga tggtgatgtc aacggtcata agttttccgt
     1441 gcgtggcgag ggtgaaggtg acgcaactaa tggtaaactg acgctgaagt tcatctgtac
     1501 tactggtaaa ctgccggtac cttggccgac tctggtaacg acgctgactt atggtgttca
     1561 gtgctttgct cgttatccgg accatatgaa gcagcatgac ttcttcaagt ccgccatgcc
     1621 ggaaggctat gtgcaggaac gcacgatttc ctttaaggat gacggcacgt acaaaacgcg
     1681 tgcggaagtg aaatttgaag gcgataccct ggtaaaccgc attgagctga aaggcattga
     1741 ctttaaagaa gacggcaata tcctgggcca taagctggaa tacaatttta acagccacaa
     1801 tgtttacatc accgccgata aacaaaaaaa tggcattaaa gcgaatttta aaattcgcca
     1861 caacgtggag gatggcagcg tgcagctggc tgatcactac cagcaaaaca ctccaatcgg
     1921 tgatggtcct gttctgctgc cagacaatca ctatctgagc acgcaaagcg ttctgtctaa
     1981 agatccgaac gagaaacgcg atcatatggt tctgctggag ttcgtaaccg cagcgggcat
     2041 cacgcatggt atggatgaac tgtacaaatg atgataataa tctagaccag gcatcaaata
     2101 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     2161 gctctctact agagtcacac tggctcacct tcgggtgggc ctttctgcgt ttataggtac
     2221 ccgtggatcc tctagagtcg acctgcaggc atgcaagctt tttacggcta gctcagtcct
     2281 aggtatagtg ctagcccagc cagagaaaca ctctttaaca gggggtagta tgatgtctcg
     2341 tttagataaa agtaaagtga ttaacagcgc attagagctg cttaatgagg tcggaatcga
     2401 aggtttaaca acccgtaaac tcgcccagaa gctaggtgta gagcagccta cattgtattg
     2461 gcatgtaaaa aataagcggg ctttgctcga cgccttagcc attgagatgt tagataggca
     2521 ccatactcac ttttgccctt tagaagggga aagctggcaa gattttttac gtaataacgc
     2581 taaaagtttt agatgtgctt tactaagtca tcgcgatgga gcaaaagtac atttaggtac
     2641 acggcctaca gaaaaacagt atgaaactct cgaaaatcaa ttagcctttt tatgccaaca
     2701 aggtttttca ctagagaatg cattatatgc actcagcgca gtggggcatt ttactttagg
     2761 ttgcgtattg gaagatcaag agcatcaagt cgctaaagaa gaaagggaaa cacctactac
     2821 tgatagtatg ccgccattat tacgacaagc tatcgaatta tttgatcacc aaggtgcaga
     2881 gccagccttc ttattcggcc ttgaattgat catatgcgga ttagaaaaac aacttaaatg
     2941 tgaaagtggg tcttaaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt
     3001 cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc gccgccctag
     3061 acctagggcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     3121 tccacagaat caggggataa cgcatgcaag aacatgtgag caaaaggcca gcaaaaggcc
     3181 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     3241 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     3301 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     3361 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     3421 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3481 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3541 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3601 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
     3661 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3721 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3781 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3841 tggaacgaaa actcacgtta agggattttg gtcatgacta gtgcttggat tctcaccaat
     3901 aaaaaacgcc cggcggcaac cgagcgttct gaacaaatcc agatggagtt ctgaggtcat
     3961 tactggatct atcaacagga gtccaagcga gctcgatatc aaattacgcc ccgccctgcc
     4021 actcatcgca gtactgttgt aattcattaa gcattctgcc gacatggaag ccatcacaaa
     4081 cggcatgatg aacctgaatc gccagcggca tcagcacctt gtcgccttgc gtataatatt
     4141 tgcccatggt gaaaacgggg gcgaagaagt tgtccatatt ggccacgttt aaatcaaaac
     4201 tggtgaaact cacccaggga ttggctgaga cgaaaaacat attctcaata aaccctttag
     4261 ggaaataggc caggttttca ccataacacg ccacatcttg cgaatatatg tgtagaaact
     4321 gccggaaatc gtcgtggtat tcactccaga gcgatgaaaa cgtttcagtt tgctcatgga
     4381 aaacggtgta acaagggtga acactatccc atatcaccag ctcaccgtct ttcattgcca
     4441 tacgaaattc cggatgagca ttcatcaggc gggcaagaat gtgaataaag gccggataaa
     4501 acttgtgctt atttttcttt acggtcttta aaaaggccgt aatatccagc tgaacggtct
     4561 ggttataggt acattgagca actgactgaa atgcctcaaa atgttcttta cgatgccatt
     4621 gggatatatc aacggtggta tatccagtga tttttttctc cattttagct tccttagctc
     4681 ctgaaaatct cgataactca aaaaatacga ccggtagtga tcttatttca ttatggtgaa
     4741 agttggaacc tcttacgggc cggccaacgt ctcattttcg ccagatatcg acgtctaaga
     4801 aa
//
