LOCUS       sequence-331738-        7724 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses BhaC1 in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7724)
  AUTHORS   Daniels PN, Lee H, Splain RA, Ting CP, Zhu L, Zhao X, Moore BS, van 
            der Donk WA
  TITLE     A biosynthetic pathway to aromatic amines that uses glycyl-tRNA as 
            nitrogen donor.
  JOURNAL   Nat Chem. 2022 Jan;14(1):71-77. doi: 10.1038/s41557-021-00802-2. 
            Epub 2021 Nov 1.
  PUBMED    34725492
REFERENCE   2  (bases 1 to 7724)
  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/s41557-021-00802-2"; journalName: "Nat Chem"; date: 
            "2022-01"; volume: "14"; issue: "1"; pages: "71-77"
COMMENT     Sequence Label: pET15b-bhaC1
FEATURES             Location/Qualifiers
     source          1..7724
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by 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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2119..2136
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2146..2163
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2225..2243)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2239..2286
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(2461..2466,2467..2483,2484..2489))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(2516..2534)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2602..2706
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2707..2775,2776..3567)
                     /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     complement(2925..2944)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3738..4326
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4227..4246
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4480..4497
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4512..4651
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4577..4596)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4737..4759
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4753..4944)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6256..7338)
                     /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     7300..7319
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7339..7416)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7622..7641
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg ggcatgttat ccgaacgaac aaaactagac
      121 gttgaatggg atcgaaagaa aattccttac acgggtgatg ttgtatcaca tgttttaaaa
      181 gtcttcgatc agaaaaaatc gaaagataaa aattgttgtg aagataaagt gtgggttctc
      241 actcataaaa aagaaaatcg aattgcaatt atcgaagagt tgaaacaaca ccttgtcaac
      301 aacatagaga ctttgacgat cggaggagtt tgggaaacaa ccggttatag tggaatagct
      361 actcaattaa gaaccatcat tacttatcta ctaaaaaaag gataccacaa ttttttagaa
      421 acctttgctc cggaaatcat ttatctcttc ccagatctaa cttccgtacc accctttaga
      481 aatgctaagc aattagacgc aatagctctt tctccctcac gtcgtagatt gcataaagaa
      541 tctgaacaaa tgttcagagt cacccaaatg gtcgtaagcc ttctcttaaa atatgtagag
      601 catacaggca aagacgttgt ttttcttttt gatcaaattg ataaaatgga cgaacataca
      661 attcgttgct ttacacgcct atctaaatgt atccatcatt gtcaagctgt tgttgttgca
      721 acgttttctg aggcgtccga taatgacccg cgttttaact ttaaaataaa cactgataac
      781 cagccttaca ttagcttagc tgaaaaccgt aatcgactgc ttcatacagc acataaacag
      841 acttccccac ttgtgttaga accaacacta acaaatgatc tctttcataa aaagaatacc
      901 attatcgctt ctgagaagaa tacgcaactt atgactaaag aggttcagac gactacaact
      961 ctcctagaaa gcatacataa aagaaagtta gacaatcttg actccgaagt gattgacgct
     1021 ctagaagaaa gtatatttac tcaaaactat gaacacgcac ttttccttat caacaagacc
     1081 tccccttata tggatcatct taaaaacaaa acgaaggtag aagtatggat ttacatcggc
     1141 ataggctacg cattcatgct caagtatgag aaagctctca cattgttcca atacgccctg
     1201 aaaaattcag aagacacact gcaaaagtca gaaatacagc tacttatcgc tttgttgtat
     1261 actaagcgac tcaacaaccc actacttggc agggaaatca ttgacaacgc acttcaaaac
     1321 ataggatcac ttactggaaa tcgagtagag gtagaacgca cttggcttta taacctaaaa
     1381 gctctcacgt ttgttgagcg gcgagattta gtgaatgctt ataaaaattg tcgaaaagct
     1441 cttgagcata ttaaaaaggg cgatcgatca gaagatgcca ttcatatcaa aataaatttg
     1501 ctttccaata tttctgtttt gtatgagtac atgaataaag tagacgcatc cctaacaaaa
     1561 tggatgaaat tcgataagtt cattctacaa tctagtccag tgtttacaaa acattatttg
     1621 tatcgaaaag ctggattact ctacaaaatg aaaaactatc tcgaggcgat tgaagcctta
     1681 aaagcaagtt ataagatcgc agcagaactg cacgactatt ttcatatgga tatcatcgcc
     1741 agaggcttag gcgctattct tttcgaggaa gagaacttct ctgaagcaat tcaatggttt
     1801 agatgtagtt taaatgcaaa gtacaaatta ttagaagatg aagatattcc ccgagtagtt
     1861 gcagcaatag ctatttgttt taatcagtta ggagaaagtc acaaagggat gaaagaaatt
     1921 gaggcgtgtt tacagacaca cacaaaaggt gatgccgctc aaatggtttt ggatatgtta
     1981 actcagtggg aacatcaaga taacgacaag tggaagagct atgttaaatg ggccattcaa
     2041 aaaccagatt ctaaacttaa ccgacctttt gacttgacca atctctattg tgaaaggacg
     2101 gagagctcgt agagcagcca tcatcatcat catcacagca gcggcctggt gccgcgcggc
     2161 agccatatgc tcgaggatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct
     2221 gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt
     2281 tttttgctga aaggaggaac tatatccgga tatcccgcaa gaggcccggc agtaccggca
     2341 taaccaagcc tatgcctaca gcatccaggg tgacggtgcc gaggatgacg atgagcgcat
     2401 tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat aaactaccgc
     2461 attaaagctt atcgatgata agctgtcaaa catgagaatt cttgaagacg aaagggcctc
     2521 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     2581 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     2641 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     2701 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     2761 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     2821 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     2881 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     2941 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     3001 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     3061 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     3121 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     3181 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     3241 acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     3301 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     3361 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     3421 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     3481 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     3541 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     3601 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     3661 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     3721 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     3781 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     3841 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     3901 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     3961 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     4021 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     4081 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     4141 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     4201 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     4261 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     4321 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     4381 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     4441 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     4501 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     4561 aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc
     4621 gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc
     4681 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     4741 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta
     4801 aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag
     4861 ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag
     4921 ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg
     4981 ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca
     5041 tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca
     5101 gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca
     5161 gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt
     5221 ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt
     5281 cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg
     5341 ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat
     5401 gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc cgcgtgcggc tgctggagat
     5461 ggcggacgcg atggatatgt tctgccaagg gttggtttgc gcattcacag ttctccgcaa
     5521 gaattgattg gctccaattc ttggagtggt gaatccgtta gcgaggtgcc gccggcttcc
     5581 attcaggtcg aggtggcccg gctccatgca ccgcgacgca acgcggggag gcagacaagg
     5641 tatagggcgg cgcctacaat ccatgccaac ccgttccatg tgctcgccga ggcggcataa
     5701 atcgccgtga cgatcagcgg tccaatgatc gaagttaggc tggtaagagc cgcgagcgat
     5761 ccttgaagct gtccctgatg gtcgtcatct acctgcctgg acagcatggc ctgcaacgcg
     5821 ggcatcccga tgccgccgga agcgagaaga atcataatgg ggaaggccat ccagcctcgc
     5881 gtcgcgaacg ccagcaagac gtagcccagc gcgtcggccg ccatgccggc gataatggcc
     5941 tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc
     6001 aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc
     6061 tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag
     6121 acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg
     6181 ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca
     6241 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     6301 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt
     6361 tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag
     6421 ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt
     6481 taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatatc
     6541 cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc
     6601 gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg
     6661 aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg
     6721 agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc
     6781 cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt
     6841 accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa
     6901 taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg
     6961 atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca
     7021 ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc
     7081 gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc
     7141 aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta
     7201 attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc
     7261 ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta
     7321 taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc
     7381 cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc tctcccttat
     7441 gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg
     7501 caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc cccggccacg gggcctgcca
     7561 ccatacccac gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga tcttccccat
     7621 cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca
     7681 cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg aaat
//
