LOCUS       sequence-376770-        5719 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses an evolved glaucine biosensor in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5719)
  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 5719)
  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-GLAU4
FEATURES             Location/Qualifiers
     source          1..5719
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        133..167
                     /label=BBa_J23114
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.1"
     misc_RNA        188..237
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             1924..2115
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2109..2131)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2272..2291
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2366..2383)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2537..3123)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2616..2635)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3294..4085,4086..4154))
                     /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(4155..4226)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(4414..5496)
                     /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     5458..5477
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5497..5574)
                     /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."
ORIGIN
        1 gagcgggaaa ccgctcccct tttttcgttt tggtcccaat ctatcgattg tatggacttc
       61 atcttcatta cctcgtatca ttgtacacct gccgaacgca agggcatggg ctgtcgacct
      121 ttgaaaagta cctttatggc tagctcagtc ctaggtacaa tgctagcaat caagatactg
      181 agcacagctg tcaccggatg tgctttccgg tctgatgagt ccgtgaggac gaaacagcct
      241 ctacaaataa ttttgtttaa actagtgaac cacgaggcct acatatggtt gctcgcccaa
      301 agtctgagga caaaaagcag gcattgcttg aagcggcaac tcaagccatc gcgcaatcag
      361 gcattgccgc tagtaccgct gtaattgcac gcaatgcggg agttgcggaa gggacgttgt
      421 tccgctattt cgcaacgaaa gatgagttga tcaacaccct ttacttacat ttgacccagg
      481 actggtgcca atcaatcatc atggaattgg atcgttctat tactgacgct aagatgatga
      541 cccgtttttt gtggaacagt tggattagct ggggattgaa ccacccagct cgccatcgtg
      601 ccattcgtca gttggcggtt tctgaaaagt tgacgaagga aaccgaacaa cgcgcggatg
      661 atatgttccc ggagttacgc gaccacctgc accgtaacgt tcttatggtg tttatgtccg
      721 acgagtaccg cgccttcggc gacgggttgt tcttggcgct tgctgagacg actatggatt
      781 tcgctgcgcg cgacccggct cgcgctggtg agtacattgc gttgggcttc gaggctatgt
      841 ggcgcgcatt ggcccgcgaa gagcagtaat aatcctaaga attcgaggag tgcaggctcg
      901 gtaacatacg gtctagctat ctgactatcg ccgctgtgag ctcggtacca aattccagaa
      961 aagaggccgc gaaagcggcc tttttcgttt tggtccaagc ccgatgcgcc agagttgttt
     1021 ctgaaacatg gcaaaggtat cactagtctt cgcggccgcc atgctgtcca ggcaggtaga
     1081 tgacgaccat cagggacagc ttcaaggatc gctcgcggct cttaccagcc taacttcgat
     1141 cattggaccg ctgatcgtca cggcgattta tgccgcctcg gcgagcacat ggaacgggtt
     1201 ggcatggatt gtaggcgccg ccctatacct tgtctgcctc cccgcgttgc gtcgcggtgc
     1261 atggagccgg gccacctcga cctgaatgga agccggcggc acctcgctaa cggattcacc
     1321 actccaagaa ttggagccaa tcaattcttg cggagaactg tgaatgcgca aaccaaccct
     1381 tggcagaaca tatccatcgc gtccgccatc tccagcagcc gcacgcggcg catctcgggc
     1441 agcgttgggt cctggccacg ggtgcgcatg atcgtgctcc tgtcgttgag gacccggcta
     1501 ggctggcggg gttgccttac tggttagcag aatgaatcac cgatacgcga gcgaacgtga
     1561 agtgactgct gctgcaaaac gtctgcgacc tgagcaacaa catgaatggt catcggtttc
     1621 cgtgtttcgt aaagtctgga aacgcggaag tcagcgccct gcaccattat gttccggatc
     1681 tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt aacgaagcgc
     1741 tggcattgac cctgagtgat ttttctctgg tcccgccgca tccataccgc cagttgttta
     1801 ccctcacaac gttccagtaa ccgggcatgt tcatcatcag taacccgtat cgtgagcatc
     1861 ctctctcgtt tcatcggtat cattaccccc atgaacagaa atccccctta cacggaggca
     1921 tcagtgacca aacaggaaaa aaccgccctt aacatggccc gctttatcag aagccagaca
     1981 ttaacgcttc tggagaaact caacgagctg gacgcggatg aacaggcaga catctgtgaa
     2041 tcgcttcacg accacgctga tgagctttac cgcagctgcc tcgcgcgttt cggtgatgac
     2101 ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat
     2161 gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca
     2221 gccatgaccc agtcacgtag cgatagcgga gtgtatactg gcttaactat gcggcatcag
     2281 agcagattgt actgagagtg caccggtgtg aaataccgca cagatgcgta aggagaaaat
     2341 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     2401 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     2461 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     2521 ccgcgttgct ggcgtttttc cataggctcc gccccctgac gagcatcaca aaaatcgacg
     2581 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     2641 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     2701 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     2761 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     2821 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     2881 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     2941 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     3001 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     3061 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaggatct
     3121 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     3181 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     3241 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctaa cagttaccaa
     3301 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     3361 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     3421 gcaatgatac cgcgcgatcc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     3481 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     3541 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     3601 gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     3661 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     3721 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     3781 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     3841 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     3901 ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     3961 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     4021 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     4081 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     4141 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     4201 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata tgcgccttga gcgacacgaa
     4261 ttatgcagtg atttacgacc tgcacagcca taccacagct tccgatggct gcctgacgcc
     4321 agaagcattg gtgcaccgtg cagtcgatga taagctgtca aacatgagaa ttgtgcctaa
     4381 tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     4441 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     4501 gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac
     4561 cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa
     4621 atcctgtttg atggtggtta acggcgggat ataacatgag ctatcttcgg tatcgtcgta
     4681 tcccactacc gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc
     4741 gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag
     4801 catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat
     4861 cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga
     4921 gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg
     4981 ctccacgccc agtcgcgtac catcttcatg ggagaaaata atactgttga tgggtgtctg
     5041 gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc
     5101 atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt
     5161 gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct
     5221 ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag
     5281 ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc
     5341 cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt
     5401 cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc
     5461 atactctgcg acatcgtata acgttactgg tttcacattc accaccctga attgactctc
     5521 ttccgggcgc tatcatgcca taccgcgaaa ggttttgcac cattcgatgg tgtcgggacg
     5581 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgttgcgg ccgcgaagac
     5641 agcgttatca gagatgagac accgtgcgat aatgtcgggc aatcaggtgc gactcggtac
     5701 caaattccag aaaagaggg
//
