LOCUS       sequence-430355-        8418 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  S. aureus USA300 GyrA Codon-optimized for E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8418)
  AUTHORS   French S, Guo ABY, Ellis MJ, Deisinger JP, Johnson JW, Rachwalski K,
            Piquette ZA, Lluka T, Zary M, Gamage S, Magolan J, Brown ED
  TITLE     A platform for predicting mechanism of action based on bacterial 
            transcriptional responses identifies an unusual DNA gyrase 
            inhibitor.
  JOURNAL   Cell Rep. 2024 Apr 4;43(4):114053. doi: 
            10.1016/j.celrep.2024.114053.
  PUBMED    38578824
REFERENCE   2  (bases 1 to 8418)
  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: "Cell Rep.";
            date: "2024-04-4"; volume: "43(4):114053. doi"; pages: " 
            10.1016/j.celrep.2024.114053"
COMMENT     Sequence Label: pET151-GyrA
FEATURES             Location/Qualifiers
     source          1..8418
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        45..122
                     /gene="lacI"
                     /label=lacI promoter
     CDS             123..1205
                     /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     complement(142..161)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2517..2708
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2702..2724)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2810..2949
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2865..2884
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2964..2981)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3135..3723)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3215..3234)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3894..4685,4686..4754))
                     /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     4517..4536
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4755..4859)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4927..4945
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(5183..5230)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     5226..5244
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(7983..8003)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(8004..8045)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(8046..8063)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             8074..8096
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    8111..8135
                     /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)."
     primer_bind     complement(8135..8154)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8136..8154)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8238..8257)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 cttcctaatg caggagtcgc ataagggaga gcgtcgagat cccggacacc atcgaatggc
       61 gcaaaacctt tcgcggtatg gcatgatagc gcccggaaga gagtcaattc agggtggtga
      121 atgtgaaacc agtaacgtta tacgatgtcg cagagtatgc cggtgtctct tatcagaccg
      181 tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa aacgcgggaa aaagtggaag
      241 cggcgatggc ggagctgaat tacattccca accgcgtggc acaacaactg gcgggcaaac
      301 agtcgttgct gattggcgtt gccacctcca gtctggccct gcacgcgccg tcgcaaattg
      361 tcgcggcgat taaatctcgc gccgatcaac tgggtgccag cgtggtggtg tcgatggtag
      421 aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa tcttctcgcg caacgcgtca
      481 gtgggctgat cattaactat ccgctggatg accaggatgc cattgctgtg gaagctgcct
      541 gcactaatgt tccggcgtta tttcttgatg tctctgacca gacacccatc aacagtatta
      601 ttttctccca tgaagacggt acgcgactgg gcgtggagca tctggtcgca ttgggtcacc
      661 agcaaatcgc gctgttagcg ggcccattaa gttctgtctc ggcgcgtctg cgtctggctg
      721 gctggcataa atatctcact cgcaatcaaa ttcagccgat agcggaacgg gaaggcgact
      781 ggagtgccat gtccggtttt caacaaacca tgcaaatgct gaatgagggc atcgttccca
      841 ctgcgatgct ggttgccaac gatcagatgg cgctgggcgc aatgcgcgcc attaccgagt
      901 ccgggctgcg cgttggtgcg gatatctcgg tagtgggata cgacgatacc gaagacagct
      961 catgttatat cccgccgtta accaccatca aacaggattt tcgcctgctg gggcaaacca
     1021 gcgtggaccg cttgctgcaa ctctctcagg gccaggcggt gaagggcaat cagctgttgc
     1081 ccgtctcact ggtgaaaaga aaaaccaccc tggcgcccaa tacgcaaacc gcctctcccc
     1141 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     1201 agtgagcgca acgcaattaa tgtaagttag ctcactcatt aggcaccggg atctcgaccg
     1261 atgcccttga gagccttcaa cccagtcagc tccttccggt gggcgcgggg catgactatc
     1321 gtcgccgcac ttatgactgt cttctttatc atgcaactcg taggacaggt gccggcagcg
     1381 ctctgggtca ttttcggcga ggaccgcttt cgctggagcg cgacgatgat cggcctgtcg
     1441 cttgcggtat tcggaatctt gcacgccctc gctcaagcct tcgtcactgg tcccgccacc
     1501 aaacgtttcg gcgagaagca ggccattatc gccggcatgg cggccgacgc gctgggctac
     1561 gtcttgctgg cgttcgcgac gcgaggctgg atggccttcc ccattatgat tcttctcgct
     1621 tccggcggca tcgggatgcc cgcgttgcag gccatgctgt ccaggcaggt agatgacgac
     1681 catcagggac agcttcaagg atcgctcgcg gctcttacca gcctaacttc gatcattgga
     1741 ccgctgatcg tcacggcgat ttatgccgcc tcggcgagca catggaacgg gttggcatgg
     1801 attgtaggcg ccgccctata ccttgtctgc ctccccgcgt tgcgtcgcgg tgcatggagc
     1861 cgggccacct cgacctgaat ggaagccggc ggcacctcgc taacggattc accactccaa
     1921 gaattggagc caatcaattc ttgcggagaa ctgtgaatgc gcaaaccaac ccttggcaga
     1981 acatatccat cgcgtccgcc atctccagca gccgcacgcg gcgcatctcg ggcagcgttg
     2041 ggtcctggcc acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc
     2101 ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact
     2161 gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt
     2221 cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg atctgcatcg
     2281 caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag cgctggcatt
     2341 gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac
     2401 aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc
     2461 gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga
     2521 ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc
     2581 ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc
     2641 acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat gacggtgaaa
     2701 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     2761 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga
     2821 cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat
     2881 tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     2941 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     3001 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     3061 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     3121 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     3181 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     3241 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     3301 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     3361 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     3421 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     3481 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     3541 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     3601 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     3661 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     3721 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     3781 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     3841 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     3901 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     3961 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     4021 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     4081 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     4141 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     4201 gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     4261 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     4321 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     4381 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     4441 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     4501 ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     4561 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     4621 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     4681 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     4741 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     4801 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     4861 acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc
     4921 tataaaaata ggcgtatcac gaggcccttt cgtcttcaag aattaattct catgtttgac
     4981 agcttatcat cgataagcta gctttaatgc ggtagtttat cacagttaaa ttgctaacgc
     5041 agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg caccgtcacc
     5101 ctggatgctg taggcatagg cttggttatg ccggtactgc cgggcctctt gcgggatatc
     5161 cggatatagt tcctcctttc agcaaaaaac ccctcaagac ccgtttagag gccccaaggg
     5221 gttatgctag ttattgctca gcggtggcag cagccaactc agcttccttt cgggctttgt
     5281 tagcagccgg atctgagctc gcccttttct tcatcactgc tctgctgaat atcctcttca
     5341 acacgatcca tgaaatcctg acgcacttca atacgaccat cttcatcatt ttcttcgcta
     5401 tcaataactt cggtatgaat ggcattaccc ggtgtttcat cattcacaac tgcttcacgc
     5461 tgctgttcgg tgccatcttc gctaacggtg ctggtgctct gttcatcttc gttggtttcg
     5521 tcctctgcat cttctttaac ttttgcaacg gtgctaacaa actggtcatc acccagacga
     5581 atcagacgaa caccctgtgc tgcacgacca ttctgactaa tatctgccac atccaggcga
     5641 ataataacac cggcattggt aacaatcatc aggtcctctt caccggtaac ggtggtaatg
     5701 caaacaacat taccattacg ttcggtaatg gttgcggttt taatgccttt accaccacga
     5761 ttgctcagac gataatcatt caccggtgta cgtttaccat aaccattttc ggtcacaacc
     5821 agcacttcat caacgctatt tgcatgtgca acatccagac caacaacttc atcaccttcg
     5881 cgcagggtaa tacctttaac accggttgcg gtacgaccca gcggacgcag ggtgctttca
     5941 ggaaaacgaa tcaggcttgc atggctggtg ccaatcagaa tatcttcttg accgctggtc
     6001 agacgcactg cgatcagctc atcatcttca cgaaagctaa tggcgatttt gccattgcga
     6061 ttaatgcgag aaaaattgct cagtgcgcta cgtttaacaa caccacgttt ggttgcaaac
     6121 accagaaagt tatcttcgct ttccagatct ttaacggcaa tcatggtgct aatcacttca
     6181 tcgttttcca gctcaattgc attaacaacc ggaatacctt tgctctgacg gctcagttcc
     6241 ggcacttcat aacctttcag tttatacacg cgacctttat tggtgaaaaa cagaacgtga
     6301 tcatgggtgc tcagggtaac cagctggcta acaaaatctt cttccagggt attcatgccc
     6361 tgaacaccac gaccaccgcg attctgtgca cgataggtgc taaccggcag acgtttaatg
     6421 tagttgttgt ggctcagcgt aataacaatt tgttcttccg gaatcagatc ctcatcttcc
     6481 aggttttcaa aaccacctaa ctgaatttcc gtacgacgat catcaccaaa acgatcacga
     6541 atttctgtca gttcatcacg taccagctgc agcagaactt cttcatctgc cagaatggct
     6601 tccagttcgc tgatatagtt cagcagctca ttatattctg cttcaatttt gtcgcgttcc
     6661 agaccggtca gacgacgcag acgcatatcc agaattgcct gtgcctgttt ttcgctcagt
     6721 ttaaaacgct gctgcaggct ttccattgca actttatcgg tatcgctttc gcgaatggtg
     6781 ctgataattt catcaatatg atccagggca atacgcagac cttccagaat atgggcacga
     6841 tcttttgctt tacgcagatt atactgggta cgacgacgaa caacggtttt ctgatgttcc
     6901 agatagtgaa ccagtgcttc tttcagattg atcagtttcg gacgaccatt aaccagtgca
     6961 atcatattaa caccaaagct ggtctgcagc ggtgtctgtt tatacaggtt gttcagaatc
     7021 acgctggcat ttgcatcttt acgaacatca ataacaacac gaacaccggt acgcaggctg
     7081 gtttcatcac gcagatcggt aataccatca atctttttat cgcgaaccag ttcggcaatt
     7141 ttctcaatca tacgggcttt gttaacctga aacggaattt cggtaacaac aatacgctga
     7201 cgaccaccac cacgttcttc aataactgca cggctacgca tctgaatgct accacgacct
     7261 gtttcatatg cacgacgaat accactttta cccagaatca gacctgcggt cggaaaatca
     7321 ggaccttcaa tatcttccat cagttctgca atgctaatat ccggattttt gctcaggctc
     7381 agcacaccat taatcagttc ggtcagatta tgaggcggaa tattggttgc cataccaact
     7441 gcaataccgc ttgcaccatt tgccagcaga ttcggaaaac gtgcaggcag aacgctcggt
     7501 tcacgttcat tgccatcata gttgtcgatg aaatcgatgg tatctttgtt aatatcacgc
     7561 agcagttcca gggtgatttt tgtcatacgt gcttcggtat aacgcattgc tgctgcacca
     7621 tcaccatcca tgctaccaaa attaccctga ccatcaacca gcggataacg atagctaaaa
     7681 tcctgtgcca tacgaaccat tgcttcataa atgctcagat caccatgcgg atgatattta
     7741 cccatcacat caccgacaat acgtgcgctc tttttatagc ttttatccgg tgtcataccc
     7801 tgttcattca ggccatacag aatacgacga tgaaccggtt tcagaccatc acgaacatcc
     7861 ggcagggcac gtgcaacaat aacgctcatt gcataatcca gaaaggattc acgcatttcg
     7921 ctggtaatat tgcgttcatt aatacggctc tgcggcagtt ctgccatggt gaagggatca
     7981 attccctgaa aatacaggtt ttccgtagaa tcgagaccga ggagagggtt agggataggc
     8041 ttaccatggt gatggtgatg atgcatatgt atatctcctt cttaaagtta aacaaaatta
     8101 tttctagagg ggaattgtta tccgctcaca attcccctat agtgagtcgt attaatttcg
     8161 cgggatcgag atctcgatcc tctacgccgg acgcatcgtg gccggcatca ccggcgccac
     8221 aggtgcggtt gctggcgcct atatcgccga catcaccgat ggggaagatc gggctcgcca
     8281 cttcgggctc atgagcgctt gtttcggcgt gggtatggtg gcaggccccg tggccggggg
     8341 actgttgggc gccatctcct tgcatgcacc attccttgcg gcggcggtgc tcaacggcct
     8401 caacctacta ctgggctg
//
