LOCUS       sequence-455756-        8486 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses precursor peptide and lanthipeptide synthetase from LanIII
            (FAST-RiPPs) in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8486)
  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 8486)
  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
FEATURES             Location/Qualifiers
     source          1..8486
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      2621..2668
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2709..3164
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2796..2815)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3006..3027
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3256..4071)
                     /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(3256..4071)
                     /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     3979..3998
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4063..4082
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4193..4781
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4682..4701
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4935..4952
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4967..5106
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5032..5051)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5192..5214
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5208..5399)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6208..7290)
                     /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     7252..7271
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7291..7368)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7574..7593
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        7681..7699
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7700..7724
                     /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             7741..7758
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      8069..8116
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        8121..8139
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    8140..8164
                     /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)."
ORIGIN
        1 tacctggatc tggtaaagcc gcatcttgat gcaggctgga aagttattcc gggcgggtat
       61 tggacccgtt gttggccgaa ggaagggcat ttagctcatg gttggaagat tcacttagca
      121 gcgtccaatg aaaccgcagt ggaattactg caacgcaccc tgccattatt aacccaagcg
      181 cgtgtctctt ttaaattttg tgccgatccc gcaattctgc gcctttcctt gaacaagaac
      241 gcgccccgta caggcgctgg taaattcgtg acgatttacc cggaaaacga ggaacatttt
      301 ctgtcattgg ccgaatcgct gcaccaggcg actgagggtc tttggggccc gttcctgctt
      361 acagatcgcc catacaaagg ctcaaaggta gtgttttatc gctacggcac ccatttagcc
      421 gaggctcgcc ttaacctgcg tggagagcgc gtcccgggta tgacaggacc ggacggctct
      481 tggatggacg atcatcgcaa accgcatttc ttcttaccga aatgggcaaa agatccgtta
      541 agcggatggc gtatccgtga acaggcacca agttccggag acgtgcgtct gaaccagcgc
      601 tatcgcatcg aacgcgcttt gaaatttaat gccagcggcg gtatttatca ggcacttgac
      661 acgaaaactg ggcagaccgt gattctgcgt gaggcccgtc ccctgacctc gggctcaccc
      721 gagcgcgacg aaggcttcat tctgctggag aaagaggcac gtattctgca acgcttagag
      781 tccacggggt atactcctcg ctttgtcgaa ctctttaaac actgggaaca cctcttcttg
      841 gtacaggaga aaattgaagc cgaatcgctt tggggctacg caatcggctt ttcacgcgga
      901 tcgtcggact tgcgcccccg tgacttcttc tcacgtgtac gtgatacgat gacccagtta
      961 gccacggcac tcaaagtgat tcatgcccgc ggcattgtaa tgcgtgactt gacgcgtaat
     1021 aacattttat tcaccagtga acgccgcctg cgctttattg atctcgaatt tgcttttgaa
     1081 ttagatggag atacaccctt cacccgctcc tggactgatg gacattcgtc accgcaacaa
     1141 cgtcgttggg aacgtcctgc tccgtcggac gactggtaca cactgggcgc tatcctcatc
     1201 gaccagatcg cttttctggc gcctggcttg gacttgaacc gcgaaggcac acttcgttct
     1261 ttccgccaga cactggaaga ttatcatctg ccactgaaat tgcatgagtt agcgcttgga
     1321 ctgttggaac cggacgcgag ccgtcgttgg gatttagacc gtgtcctggc agtactgggc
     1381 gatattcccg aacccaccca tgatactcca attgttccga ttggcggaac cacgccccct
     1441 gccccaccgc cggcgccgac actcgatgta gaattggatc tcactttggc aggcatcgag
     1501 cgcttcattg tatcgaaaac cgacgatact cgtgatgatc gtttatggcc agatcgcggc
     1561 ggtagtttct ctacaaatcc actgaacctg gaattcggtg cggcgggcat tgctgcgttc
     1621 cttcatcgcc gtccaggtgg ggtgccgacc gcggctgtgg aatggatgaa acgccagtta
     1681 ccgcgtcatc gctgtccgcc gggcttgttt caagggctgg ctggtaccgc tttggttttg
     1741 caggaactgg gatgcactga acaagcccgt gccgcaatgg acgaagccgc ctcttcaagc
     1801 ctgctgcatc aggaattcga tttgttttgg ggcgactccg gttggggcct cgcaaacctg
     1861 catttctggc gccgcactgg tgaagaacag tatcttaccc gcgcattaga agtagcagaa
     1921 ttcctgcaac gcactaaaca agaaagctcg cagggtgttt gctggtcctc accaaccgaa
     1981 acgccgctgg gcctgttcca cggccaatcg ggcccagcat tattcctgac gtatttaagt
     2041 ttagcccgcc caaaagaagg tttcctggag ttggccgtca aagcactgga ttttgaagtt
     2101 gcacattgtc agtactcagg tgacgttctt ctgtggtatc cacacaccgc tgcgggtaac
     2161 gggcagccaa aatcaccaca tatgcgccac ggcacggcgg gcgtggggtc ggctttactg
     2221 cgtgcgcatg tagcgacagg tgaaccgcgt ttactgcgtt tagcagacca gtgcgcgaac
     2281 accgtgcgcg accgccacac caacaaatta tggctcaatt acggtctgtc gggttacggt
     2341 gaattcctgg tagatatgca ccgcctgctc ggcgatgaac gctatctgca ctgggcgtgc
     2401 tttctgtcgg aagccattat ggcccaccgt atccagaaag aagaagggat tgcattcccg
     2461 gcggatagtc tgctgcgtat cagttgtgac ctcggtggtg gtatggcagg aattggcttg
     2521 ttcctgcatc gtctgcgcaa tccgcgcgaa cctcattttc tgtttaatga tgaactgttg
     2581 ggtttaaatg ctattcgcgg tcagagtacc ccctaacggt ctagcataac cccttggggc
     2641 ctctaaacgg gtcttgaggg gttttttgag cgctgaaagg aggaactata tccggattgg
     2701 cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     2761 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     2821 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     2881 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     2941 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     3001 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     3061 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     3121 aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg
     3181 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3241 ctcatgaatt aattcttaga aaaactcatc gagcatcaaa tgaaactgca atttattcat
     3301 atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc
     3361 accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc
     3421 aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc
     3481 accatgagtg acgactgaat ccggtgagaa tggcaaaagt ttatgcattt ctttccagac
     3541 ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt
     3601 attcattcgt gattgcgcct gagcgagacg aaatacgcga tcgctgttaa aaggacaatt
     3661 acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc
     3721 acctgaatca ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt
     3781 gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa
     3841 ttccgtcagc cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt
     3901 gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc
     3961 acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt
     4021 ggaatttaat cgcggcctag agcaagacgt ttcccgttga atatggctca taacacccct
     4081 tgtattactg tttatgtaag cagacagttt tattgttcat gaccaaaatc ccttaacgtg
     4141 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     4201 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     4261 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4321 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4381 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     4441 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     4501 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4561 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     4621 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     4681 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     4741 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     4801 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     4861 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     4921 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt
     4981 ttctccttac gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg
     5041 ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg
     5101 gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     5161 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     5221 ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc
     5281 gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt
     5341 aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact
     5401 gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag
     5461 aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag
     5521 ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc
     5581 cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg
     5641 cagatccgga acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca
     5701 cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg
     5761 cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag
     5821 cctagccggg tcctcaacga caggagcacg atcatgcgca cccgtggggc cgccatgccg
     5881 gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga
     5941 gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag
     6001 cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc
     6061 atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag
     6121 gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg
     6181 agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     6241 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     6301 cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg
     6361 gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg
     6421 tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac
     6481 taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag
     6541 cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg
     6601 catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg
     6661 aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga
     6721 acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac
     6781 gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga
     6841 gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg
     6901 gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac
     6961 cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc
     7021 cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag
     7081 actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg
     7141 gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga
     7201 aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc
     7261 tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg
     7321 gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac
     7381 gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga
     7441 gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca
     7501 cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc
     7561 gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg
     7621 tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg cgaaatatgg
     7681 taatacgact cactataggg gaattgtgag cggataacaa ttccaaggag atatacaatg
     7741 caccaccacc accaccacgc aatgtcaaac aaaccaacgc atgtagatgt ggagctcaac
     7801 gatgcgcagc tcgatcaagt agtgggcggt caggaagttt tatcgctcca gatgatggat
     7861 gtgggtcagg cagaagttcc taactgtgtg tcatcggtgt cttgcaatag ttctgcaagc
     7921 tgcgaatcga gcgcaagcgc cgtcgtaagc gccgccacgt aacagactag ttggaagtgt
     7981 gtctagatct tagttttcgt cactagaggg cccacgcttt atttttatga tccattgatc
     8041 tcccagacgc tgcaagattt gcaacggtct agcataaccc cttggggcct ctaaacgggt
     8101 cttgaggggt tttttgccct taatacgact cactataggg gaattgtgag cggataacaa
     8161 ttccaaggag atatacaatg gctgcttggc ttcgtcctgg tgccccgccc gcaaactcca
     8221 atactccgcg ctgcacggct ggtacagcat ggagtcattc gtcactgatg tgtccaggca
     8281 tcggaagctt tcatacgcaa ccgttcctgc cttcgagtgg aacaattcgc ccgccgttga
     8341 atcacaatca tcctgcaggt cctgcccgcc acccgcgcga tagcctgcag gcacctgcct
     8401 ttctgccgcc gcgtacgtcc atgttcctga cctttctgtc acctgagtac tatgaaacac
     8461 tggaacgtca tcgtccgagc gctgaa
//
