LOCUS       sequence-381129-        5722 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 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-GLAU4
FEATURES             Location/Qualifiers
     source          1..5722
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        134..168
                     /label=BBa_J23114
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.1"
     misc_RNA        189..238
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             1926..2117
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2111..2133)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2274..2293
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2368..2385)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2539..3127)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2619..2638)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3298..4089,4090..4158))
                     /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(4159..4230)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(4418..5500)
                     /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     5462..5481
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5501..5578)
                     /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 ggagcgggaa accgctcccc ttttttcgtt ttggtcccaa tctatcgatt gtatggactt
       61 catcttcatt acctcgtatc attgtacacc tgccgaacgc aagggcatgg gctgtcgacc
      121 tttgaaaagt acctttatgg ctagctcagt cctaggtaca atgctagcaa tcaagatact
      181 gagcacagct gtcaccggat gtgctttccg gtctgatgag tccgtgagga cgaaacagcc
      241 tctacaaata attttgttta aactagtgaa ccacgaggcc tacatatggt tgctcgccca
      301 aagtctgagg acaaaaagca ggcattgctt gaagcggcaa ctcaagccat cgcgcaatca
      361 ggcattgccg ctagtaccgc tgtaattgca cgcaatgcgg gagttgcgga agggacgttg
      421 ttccgctatt tcgcaacgaa agatgagttg atcaacaccc tttacttaca tttgacccag
      481 gactggtgcc aatcaatcat catggaattg gatcgttcta ttactgacgc taagatgatg
      541 acccgttttt tgtggaacag ttggattagc tggggattga accacccagc tcgccatcgt
      601 gccattcgtc agttggcggt ttctgaaaag ttgacgaagg aaaccgaaca acgcgcggat
      661 gatatgttcc cggagttacg cgaccacctg caccgtaacg ttcttatggt gtttatgtcc
      721 gacgagtacc gcgccttcgg cgacgggttg ttcttggcgc ttgctgagac gactatggat
      781 ttcgctgcgc gcgacccggc tcgcgctggt gagtacattg cgttgggctt cgaggctatg
      841 tggcgcgcat tggcccgcga agagcagtaa taatcctaag aattcgagga gtgcaggctc
      901 ggtaacatac ggtctagcta tctgactatc gccgctgtga gctcggtacc aaattccaga
      961 aaagaggccg cgaaagcggc cttttttcgt tttggtccaa gcccgatgcg ccagagttgt
     1021 ttctgaaaca tggcaaaggt atcactagtc ttcgcggccg ccatgctgtc caggcaggta
     1081 gatgacgacc atcagggaca gcttcaagga tcgctcgcgg ctcttaccag cctaacttcg
     1141 atcattggac cgctgatcgt cacggcgatt tatgccgcct cggcgagcac atggaacggg
     1201 ttggcatgga ttgtaggcgc cgccctatac cttgtctgcc tccccgcgtt gcgtcgcggt
     1261 gcatggagcc gggccacctc gacctgaatg gaagccggcg gcacctcgct aacggattca
     1321 ccactccaag aattggagcc aatcaattct tgcggagaac tgtgaatgcg caaaccaacc
     1381 cttggcagaa catatccatc gcgtccgcca tctccagcag ccgcacgcgg cgcatctcgg
     1441 gcagcgttgg gtcctggcca cgggtgcgca tgatcgtgct cctgtcgttg aggacccggc
     1501 taggctggcg gggttgcctt actggttagc agaatgaatc accgatacgc gagcgaacgt
     1561 gaagtgactg ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg gtcatcggtt
     1621 tccgtgtttc gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga
     1681 tctgcatcgc aggatgctgc tggctaccct gtggaacacc tacatctgta ttaacgaagc
     1741 gctggcattg accctgagtg atttttctct ggtcccgccg catccatacc gccagttgtt
     1801 taccctcaca acgttccagt aaccgggcat gttcatcatc agtaacccgt atcgtgagca
     1861 tcctctctcg tttcatcggt atcattaccc ccatgaacag aaatccccct tacacggagg
     1921 catcagtgac caaacaggaa aaaaccgccc ttaacatggc ccgctttatc agaagccaga
     1981 cattaacgct tctggagaaa ctcaacgagc tggacgcgga tgaacaggca gacatctgtg
     2041 aatcgcttca cgaccacgct gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg
     2101 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     2161 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg
     2221 cagccatgac ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc
     2281 agagcagatt gtactgagag tgcaccggtg tgaaataccg cacagatgcg taaggagaaa
     2341 ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     2401 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     2461 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     2521 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     2581 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     2641 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     2701 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     2761 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     2821 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     2881 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     2941 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     3001 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3061 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3121 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3181 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     3241 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctaacagtta
     3301 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     3361 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     3421 tgctgcaatg ataccgcgcg atccacgctc accggctcca gatttatcag caataaacca
     3481 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     3541 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     3601 tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     3661 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     3721 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     3781 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     3841 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     3901 ttgcccggcg tcaacacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     3961 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     4021 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     4081 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     4141 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     4201 ttgtctcatg agcggataca tatttgaatg tatttagaaa aatatgcgcc ttgagcgaca
     4261 cgaattatgc agtgatttac gacctgcaca gccataccac agcttccgat ggctgcctga
     4321 cgccagaagc attggtgcac cgtgcagtcg atgataagct gtcaaacatg agaattgtgc
     4381 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4441 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4501 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     4561 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     4621 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctatct tcggtatcgt
     4681 cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     4741 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     4801 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     4861 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg
     4921 ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca
     4981 gatgctccac gcccagtcgc gtaccatctt catgggagaa aataatactg ttgatgggtg
     5041 tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa
     5101 tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa
     5161 gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca
     5221 cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt
     5281 gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt
     5341 gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg
     5401 ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac
     5461 cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac
     5521 tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcaccattcg atggtgtcgg
     5581 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt gcggccgcga
     5641 agacagcgtt atcagagatg agacaccgtg cgataatgtc gggcaatcag gtgcgactcg
     5701 gtaccaaatt ccagaaaaga gg
//
