LOCUS       sequence-447437-        8165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses precursor peptide and lanthipeptide synthetase from 
            LanIII-43 (FAST-RiPPs) in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8165)
  AUTHORS   Ayikpoe RS, Shi C, Battiste AJ, Eslami SM, Ramesh S, Simon MA, 
            Bothwell IR, Lee H, Rice AJ, Ren H, Tian Q, Harris LA, Sarksian R, 
            Zhu L, Frerk AM, Precord TW, van der Donk WA, Mitchell DA, Zhao H
  TITLE     A scalable platform to discover antimicrobials of ribosomal origin.
  JOURNAL   Nat Commun. 2022 Oct 17;13(1):6135. doi: 10.1038/s41467-022-33890-w.
  PUBMED    36253467
REFERENCE   2  (bases 1 to 8165)
  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 
            Commun."; date: "2022-10-17"; volume: "13(1):6135. doi"; pages: " 
            10.1038/s41467-022-33890-w"
COMMENT     Sequence Label: pET28:LanIII-43
FEATURES             Location/Qualifiers
     source          1..8165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    775..914
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(840..859)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1000..1022
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1016..1207)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2016..3098)
                     /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     3060..3079
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3099..3176)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3382..3401
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        3489..3507
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3508..3532
                     /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)."
     CDS             3549..3566
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3877..3924
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        3929..3947
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3948..3972
                     /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)."
     terminator      6594..6641
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      6682..7137
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6769..6788)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6979..7000
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(7229..8044)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(7229..8044)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     7952..7971
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     8036..8055
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
      781 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt
      841 acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact
      901 gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
      961 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     1021 ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt
     1081 cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca
     1141 gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt
     1201 tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga
     1261 aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac
     1321 gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg
     1381 gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc
     1441 ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt
     1501 acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc
     1561 agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac
     1621 cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc cgtggggccg
     1681 ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga
     1741 aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg
     1801 cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta
     1861 cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc
     1921 accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct
     1981 aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     2041 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     2101 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
     2161 accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga
     2221 aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg
     2281 tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt
     2341 gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc
     2401 agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct
     2461 atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc
     2521 gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga
     2581 tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc
     2641 tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg
     2701 gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga
     2761 ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg
     2821 ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc
     2881 agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt
     2941 gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt
     3001 ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg
     3061 gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc
     3121 tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg
     3181 atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag
     3241 gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     3301 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     3361 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     3421 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg
     3481 aaatatggta atacgactca ctatagggga attgtgagcg gataacaatt ccaaggagat
     3541 atacaatgca ccaccaccac caccacgcaa tgtctcgctt cgatgtactt agcttacaaa
     3601 gcttaaagaa acataagaac aacgataaac cgattggctc gaacgtgagc gtaaactgca
     3661 agacccagtc cggcttgagc ctgttcttct gcgtttcgcc gaaataacta gaagcccggc
     3721 agagttagtt cgaacctaat gccacaaatg agacaggacg ccaatgggta ccggacatta
     3781 ggtcgagctc agttcggtaa cggagagacc ctgcggcgta cttaattatg catatgaaac
     3841 gcgcccaagt gacgccaggc aagtctcagc agcggtctag cataacccct tggggcctct
     3901 aaacgggtct tgaggggttt tttgccctta atacgactca ctatagggga attgtgagcg
     3961 gataacaatt ccaaggagat atacaatgga taaccgctat atcaaataca tcaataagaa
     4021 tccatattat agcgtagccg atgagatcag ccaagggctg cttaacgtgg actatccctt
     4081 cgataactac gttctgatcg atggggagca ttggacaaat ttgatgagta agagcgctcg
     4141 tctgcccaaa cagggctgga agattcatgt gtcaacggac atccgcgaat ctcaaaagac
     4201 attagacatc gttagcaaac tgatgattga acgcaacatc agctttaagt atgtgcgttc
     4261 taacaaagag ctggtcttga aagattcgaa gtacggggat cgcggttctt ccggtaagtt
     4321 tatcacgatc tatccagaaa acaacgatca attcattgat ctgcttcgcc tgctggaatc
     4381 taatctggcg catctgaaac caggacctta catcctgaat gataaacgtt ggtacaacag
     4441 taatatttac tttcgttatg gcgcctttat gtcgcgctac gtttacatcg atggagaaaa
     4501 ggtggatgcc attgaaaatc tcgaaggcga actgattgaa gataagcgcg tcccttacta
     4561 ttatctgccc aattttgtcg aagaaccgat tgagattcgt aaaatggatg aggagctgaa
     4621 ccaagtagat accagctcgc ctctggacaa tttcgacatc aatgaagcgt tgcactttag
     4681 caacggcggc ggtgtgtata tttgtaagaa caaaagcaac aacatgaagg ttattttaaa
     4741 agaaggccgc ccgcacgcag cagtggacgc gcagggtcaa gatgcgttca gccgtattgt
     4801 taacgaaagc gaaattctgg acaaactgga aaaggcgcag taccccgtta agaaaatttc
     4861 tagcttccgt gcatgggaac attattttat tgaagaggag tatatcgaag gcgacagcct
     4921 ttcagagtgg ctcgtgaaga actacccgtt tagcagcaat atcaagaatg aatcatacac
     4981 caagagctgc attaacatca ttaatcagct gatcgattct attcacgaat tgcatcgtaa
     5041 cgacattggt atgggcgatt tacagccggc gaatattatt atcacaccgg aagaaaagat
     5101 ccgtctcatc gattttgaaa ccgcaagctc aaccagtgat agtctgagtg gactgatgac
     5161 cccgggcttc atcggcaacc aggaaatgaa taaggaacag agcgactggt ttgcactgct
     5221 gcgtattgca aagcaactgt tcgtaccgat cggctgtgtt caagacatca gctggaatat
     5281 ggataccatt cactcgaatt ggattcgcgt tgtctttggt gagaaagcca acacgatcat
     5341 tgagcgcatt gagtcgttat gtgaatttta cggctcgcgt ccaatggaag aattgttgac
     5401 aacgaacggt cagatgaaac agaaatttga tctgtccgcc atgaagttaa aacttcgcaa
     5461 ctcaatcctg aaggacgtga agaatgagga gcgtttactc ccgggcgata ttcgccaata
     5521 cgagatggag actggcatgc tgaatgtatt aaccggtggt tttggtgtag cgatggcgtt
     5581 acatcgcacc ggcgggatca accagaaggc gaaagactgg gtagaaaatc aggatatcaa
     5641 taagcttatc caactcgaga atgggctgtt taccggtaat atgggtatcg cgtccgtgct
     5701 gtgggagctg ggctaccagg aaaaggccaa ggcgctgttc gacagcttca ttgacttcaa
     5761 acatatcgag gatatcagct tcgcaacggg tctgtctggc attggttttg cgtacctggg
     5821 ttttagttat gaggtgaacg atcccaagta cctgaatatt tgcctgtata ttggtgaact
     5881 gctgattaac aaactgaatg cggatgcacc gattattacc tacgactatg atgtagtgga
     5941 taagggcgtc atgacctctt gggcaggtgt ctcattatac ttcaccgccc tttacaagaa
     6001 aactaacgat aataaatggt tggttctttc tgaacaggcc cttgaaaagg agatgaaatt
     6061 aggcatcttt gatgataccg gcctttacca gattgatgat gattttcgta tccttccata
     6121 cctggccggc ggcggctctg gcctcgcgtt gccgattgtg gagctggaga ttaatgccaa
     6181 gctgaagaaa tggaataaag agattgaagg catcagtaaa attagtaaat ccaagtgctt
     6241 ttataacgcg ggtttgttcc agggcaccac cggcattttg gcggtcgcta accttttaga
     6301 gttgtacacg cagaaatcta gcttggtgaa cagcgcttta tttacactga accttcactt
     6361 attagaagat gcagactgca ttttcgtgcc gggcgattcg tgctttcgca tcagcggcga
     6421 cattatgtct ggcagttcag gtttattact tacaattaac gacattctgg aaagcaagaa
     6481 ccacagttgg attccgatct taaacctgga aaaggtgttt agcagcagca cctttacaga
     6541 agaaagttta cagaatattg ttgaaccgtt gagcagcatc gtgcgttgac ggtctagcat
     6601 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gagcgctgaa aggaggaact
     6661 atatccggat tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     6721 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     6781 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     6841 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     6901 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     6961 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     7021 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     7081 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg
     7141 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     7201 aaatatgtat ccgctcatga attaattctt agaaaaactc atcgagcatc aaatgaaact
     7261 gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt ttctgtaatg
     7321 aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat cggtctgcga
     7381 ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa ataaggttat
     7441 caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa agtttatgca
     7501 tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa tcactcgcat
     7561 caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt
     7621 taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact gccagcgcat
     7681 caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct gttttcccgg
     7741 ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc ttgatggtcg
     7801 gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta acatcattgg
     7861 caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc ccatacaatc
     7921 gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac ccatataaat
     7981 cagcatccat gttggaattt aatcgcggcc tagagcaaga cgtttcccgt tgaatatggc
     8041 tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt catgaccaaa
     8101 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     8161 tcttc
//
