LOCUS       sequence-381131-        5698 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a generalist biosensor responsive to bicyclic monoterpenes
            in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5698)
  AUTHORS   d'Oelsnitz S, Nguyen V, Alper HS, Ellington AD
  TITLE     Evolving a Generalist Biosensor for Bicyclic Monoterpenes.
  JOURNAL   ACS Synth Biol. 2022 Jan 5. doi: 10.1021/acssynbio.1c00402.
  PUBMED    34985281
REFERENCE   2  (bases 1 to 5698)
  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. 2022 Jan 5. doi: 10.1021/acssynbio.1c00402."
COMMENT     Sequence Label: pReg-FEN2
FEATURES             Location/Qualifiers
     source          1..5698
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      204..213
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1492..1683
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1677..1699)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1840..1859
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1934..1951)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2105..2693)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2185..2204)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2864..3655,3656..3724))
                     /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"
     primer_bind     3487..3506
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3725..3796)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(3984..5066)
                     /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     5028..5047
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5067..5144)
                     /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        5422..5456
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
     misc_RNA        5477..5526
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
ORIGIN
        1 ttatggcgat cttcaaggtt tacataaagc agctatcgat gaaacattcc gtcaggtcgc
       61 agaggcatat cacggcggaa ccgaggagcg tggaccgtta aaaggcatcc gcgacggctg
      121 ggccacattc ctgcagttcg cctattcaga gccaaatatg tgccgtatgt tggttcaaca
      181 tatcatggca ggagaacccc cttccatggt ggctgatacc cttcgcggag tcgcagatga
      241 tttagctcag ttccatgcac aaggccgctt aaccttccca ccgcgtgaga ctgcccaatt
      301 attatggatg ggcgctttgg gtgcactgac ctatgcactt tcacgtgaag gagcaggtta
      361 cactcaggac ttggcactgc agaaggctaa attagatatt acacttgtag cactgtttaa
      421 tatcgaggag gaataataat cctaagaatt cgaggagtgc aggctcggta acatacggtc
      481 tagctatctg actatcgccg ctgtgagctc ggtaccaaat tccagaaaag aggccgcgaa
      541 agcggccttt tttcgttttg gtccaagccc gatgcgccag agttgtttct gaaacatggc
      601 aaaggtatca ctagtcttcg cggccgccat gctgtccagg caggtagatg acgaccatca
      661 gggacagctt caaggatcgc tcgcggctct taccagccta acttcgatca ttggaccgct
      721 gatcgtcacg gcgatttatg ccgcctcggc gagcacatgg aacgggttgg catggattgt
      781 aggcgccgcc ctataccttg tctgcctccc cgcgttgcgt cgcggtgcat ggagccgggc
      841 cacctcgacc tgaatggaag ccggcggcac ctcgctaacg gattcaccac tccaagaatt
      901 ggagccaatc aattcttgcg gagaactgtg aatgcgcaaa ccaacccttg gcagaacata
      961 tccatcgcgt ccgccatctc cagcagccgc acgcggcgca tctcgggcag cgttgggtcc
     1021 tggccacggg tgcgcatgat cgtgctcctg tcgttgagga cccggctagg ctggcggggt
     1081 tgccttactg gttagcagaa tgaatcaccg atacgcgagc gaacgtgaag tgactgctgc
     1141 tgcaaaacgt ctgcgacctg agcaacaaca tgaatggtca tcggtttccg tgtttcgtaa
     1201 agtctggaaa cgcggaagtc agcgccctgc accattatgt tccggatctg catcgcagga
     1261 tgctgctggc taccctgtgg aacacctaca tctgtattaa cgaagcgctg gcattgaccc
     1321 tgagtgattt ttctctggtc ccgccgcatc cataccgcca gttgtttacc ctcacaacgt
     1381 tccagtaacc gggcatgttc atcatcagta acccgtatcg tgagcatcct ctctcgtttc
     1441 atcggtatca ttacccccat gaacagaaat cccccttaca cggaggcatc agtgaccaaa
     1501 caggaaaaaa ccgcccttaa catggcccgc tttatcagaa gccagacatt aacgcttctg
     1561 gagaaactca acgagctgga cgcggatgaa caggcagaca tctgtgaatc gcttcacgac
     1621 cacgctgatg agctttaccg cagctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc
     1681 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     1741 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag
     1801 tcacgtagcg atagcggagt gtatactggc ttaactatgc ggcatcagag cagattgtac
     1861 tgagagtgca ccggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc
     1921 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     1981 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     2041 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     2101 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     2161 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     2221 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     2281 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     2341 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     2401 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     2461 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     2521 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     2581 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     2641 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     2701 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     2761 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     2821 ttaaatcaat ctaaagtata tatgagtaaa cttggtctaa cagttaccaa tgcttaatca
     2881 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     2941 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     3001 cgcgcgatcc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     3061 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     3121 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgctg
     3181 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     3241 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     3301 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     3361 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     3421 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     3481 cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     3541 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     3601 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     3661 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     3721 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     3781 gatacatatt tgaatgtatt tagaaaaata tgcgccttga gcgacacgaa ttatgcagtg
     3841 atttacgacc tgcacagcca taccacagct tccgatggct gcctgacgcc agaagcattg
     3901 gtgcaccgtg cagtcgatga taagctgtca aacatgagaa ttgtgcctaa tgagtgagct
     3961 aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     4021 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg
     4081 gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc
     4141 tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg
     4201 atggtggtta acggcgggat ataacatgag ctatcttcgg tatcgtcgta tcccactacc
     4261 gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc
     4321 atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg
     4381 gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt
     4441 tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt
     4501 aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc
     4561 agtcgcgtac catcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca
     4621 tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca
     4681 tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc
     4741 gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt
     4801 tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg
     4861 gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg
     4921 ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg
     4981 tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg
     5041 acatcgtata acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc
     5101 tatcatgcca taccgcgaaa ggttttgcac cattcgatgg tgtcgggacg tcaggtggca
     5161 cttttcgggg aaatgtgcgc ggaaccccta tttgttgcgg ccgcgaagac agcgttatca
     5221 gagatgagac accgtgcgat aatgtcgggc aatcaggtgc gactcggtac caaattccag
     5281 aaaagagggg agcgggaaac cgctcccctt ttttcgtttt ggtcccaatc tatcgattgt
     5341 atggacttca tcttcattac ctcgtatcat tgtacacctg ccgaacgcaa gggcatgggc
     5401 tgtcgacctt tgaaaagtac ctttacggct agctcagtcc taggtatagt gctagcaatc
     5461 aagatactga gcacagctgt caccggatgt gctttccggt ctgatgagtc cgtgaggacg
     5521 aaacagcctc tacaaataat tttgtttaaa ctagctgccg cagacccgca catatggaca
     5581 ttaagcagtc gttacttcac gcagctatgc gcttacttag cgccaagggc cgcgacggag
     5641 ctaccatgcg tgcaatctgc gccgaagtgg gtgttactcc tccaactctg tatcacca
//
