LOCUS       sequence-383677-        5722 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses an evolved tetrahydropapaverine biosensor in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5722)
  AUTHORS   d'Oelsnitz S, Kim W, Burkholder NT, Javanmardi K, Thyer R, Zhang Y, 
            Alper HS, Ellington AD
  TITLE     Using fungible biosensors to evolve improved alkaloid biosyntheses.
  JOURNAL   Nat Chem Biol. 2022 Jul 7. pii: 10.1038/s41589-022-01072-w. doi: 
            10.1038/s41589-022-01072-w.
  PUBMED    35799063
REFERENCE   2  (bases 1 to 5722)
  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 Chem 
            Biol. 2022 Jul 7. pii: 10.1038/s41589-022-01072-w. doi: 
            10.1038/s41589-022-01072-w."
COMMENT     Sequence Label: pReg-THP4
FEATURES             Location/Qualifiers
     source          1..5722
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1733..1924
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1918..1940)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2081..2100
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2175..2192)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2346..2934)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2426..2445)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3105..3896,3897..3965))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(3966..4037)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(4225..5307)
                     /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"
     promoter        complement(5308..5385)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     promoter        5663..5697
                     /label=BBa_J23112
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0"
     promoter        5663..5697
                     /label=BBa_J23103
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.01"
     misc_RNA        join(5718..5722,1..45)
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
ORIGIN
        1 accggatgtg ctttccggtc tgatgagtcc gtgaggacga aacagcctct acaaataatt
       61 ttgtttaaac tagtgaacca cgaggcctac atatggttgc tcgcccaaag tctgaggaca
      121 aaaagcaggc attgcttgaa gcagcaactc aagccatcgc gcaatcaggc actgccgcta
      181 gtaccgctgt aattgcacgc aatgcgggag ttgcggaagg gacgttgttc cgctatttcg
      241 caacgaaaga tgagttgatc aacacccttt acttacattt gttccaggac tggtgccaat
      301 catccatcat ggaattggat cgttctatta ctgacgctaa gatgatgacg cgttttctct
      361 ggaacagtat cattagctgg ggattaaacc acccagctcg ccatcgtgcc attcgtcagt
      421 tggcggtttc tgaaaagttg tcgaaggaga ccgtacaacg cgcggatgat atgttcccgg
      481 agttacgcga catcgtccac cgtgaggttc ttatggtgtt tatgtccgac gagtaccgcg
      541 ccttcggcga agggttgttc ttggcgcttg ctgagacgac tatggatttc gctgcgcgcg
      601 acccggctcg cgctggtgag tacattgcgt tgggcttcga ggctatgtgg cgcgcactta
      661 cgcgcgaaga gcagtaataa tcctaagaat tcgaggagtg caggctcggt aacatacggt
      721 ctagctatct gactatcgcc gctgtgagct cggtaccaaa ttccagaaaa gaggccgcga
      781 aagcggcctt ttttcgtttt ggtccaagcc cgatgcgcca gagttgtttc tgaaacatgg
      841 caaaggtatc actagtcttc gcggccgcca tgctgtccag gcaggtagat gacgaccatc
      901 agggacagct tcaaggatcg ctcgcggctc ttaccagcct aacttcgatc attggaccgc
      961 tgatcgtcac ggcgatttat gccgcctcgg cgagcacatg gaacgggttg gcatggattg
     1021 taggcgccgc cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca tggagccggg
     1081 ccacctcgac ctgaatggaa gccggcggca cctcgctaac ggattcacca ctccaagaat
     1141 tggagccaat caattcttgc ggagaactgt gaatgcgcaa accaaccctt ggcagaacat
     1201 atccatcgcg tccgccatct ccagcagccg cacgcggcgc atctcgggca gcgttgggtc
     1261 ctggccacgg gtgcgcatga tcgtgctcct gtcgttgagg acccggctag gctggcgggg
     1321 ttgccttact ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa gtgactgctg
     1381 ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc atcggtttcc gtgtttcgta
     1441 aagtctggaa acgcggaagt cagcgccctg caccattatg ttccggatct gcatcgcagg
     1501 atgctgctgg ctaccctgtg gaacacctac atctgtatta acgaagcgct ggcattgacc
     1561 ctgagtgatt tttctctggt cccgccgcat ccataccgcc agttgtttac cctcacaacg
     1621 ttccagtaac cgggcatgtt catcatcagt aacccgtatc gtgagcatcc tctctcgttt
     1681 catcggtatc attaccccca tgaacagaaa tcccccttac acggaggcat cagtgaccaa
     1741 acaggaaaaa accgccctta acatggcccg ctttatcaga agccagacat taacgcttct
     1801 ggagaaactc aacgagctgg acgcggatga acaggcagac atctgtgaat cgcttcacga
     1861 ccacgctgat gagctttacc gcagctgcct cgcgcgtttc ggtgatgacg gtgaaaacct
     1921 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     1981 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca
     2041 gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga gcagattgta
     2101 ctgagagtgc accggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     2161 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     2221 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     2281 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     2341 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     2401 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     2461 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     2521 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     2581 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     2641 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     2701 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     2761 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     2821 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     2881 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     2941 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     3001 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     3061 tttaaatcaa tctaaagtat atatgagtaa acttggtcta acagttacca atgcttaatc
     3121 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     3181 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     3241 ccgcgcgatc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     3301 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     3361 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     3421 gcaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     3481 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     3541 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     3601 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     3661 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     3721 acacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     3781 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     3841 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     3901 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     3961 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     4021 ggatacatat ttgaatgtat ttagaaaaat atgcgccttg agcgacacga attatgcagt
     4081 gatttacgac ctgcacagcc ataccacagc ttccgatggc tgcctgacgc cagaagcatt
     4141 ggtgcaccgt gcagtcgatg ataagctgtc aaacatgaga attgtgccta atgagtgagc
     4201 taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     4261 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag
     4321 ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc
     4381 ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt
     4441 gatggtggtt aacggcggga tataacatga gctatcttcg gtatcgtcgt atcccactac
     4501 cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc
     4561 catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat
     4621 ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat
     4681 ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact
     4741 taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc
     4801 cagtcgcgta ccatcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac
     4861 atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc
     4921 atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc
     4981 cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag
     5041 ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact
     5101 ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt
     5161 gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac
     5221 gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc
     5281 gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg
     5341 ctatcatgcc ataccgcgaa aggttttgca ccattcgatg gtgtcgggac gtcaggtggc
     5401 acttttcggg gaaatgtgcg cggaacccct atttgttgcg gccgcgaaga cagcgttatc
     5461 agagatgaga caccgtgcga taatgtcggg caatcaggtg cgactcggta ccaaattcca
     5521 gaaaagaggg gagcgggaaa ccgctcccct tttttcgttt tggtcccaat ctatcgattg
     5581 tatggacttc atcttcatta cctcgtatca ttgtacacct gccgaacgca agggcatggg
     5641 ctgtcgacct ttgaaaagta ccctgatagc tagctcagtc ctagggatta tgctagcaat
     5701 caagatactg agcacagctg tc
//
