LOCUS       sequence-12883-e        6788 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6788)
  AUTHORS   Blumenthal A, Trujillo C, Ehrt S, Schnappinger D
  TITLE     Simultaneous analysis of multiple Mycobacterium tuberculosis 
            knockdown mutants in vitro and in vivo.
  JOURNAL   PLoS One. 2010 Dec 22;5(12):e15667. doi: 
            10.1371/journal.pone.0015667.
  PUBMED    21203517
REFERENCE   2  (bases 1 to 6788)
  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: "PLoS One.";
            date: "2010-12-22"; volume: "5(12):e15667. doi"; pages: " 
            10.1371/journal.pone.0015667"
COMMENT     Sequence Label: pDE43-MCSq22
FEATURES             Location/Qualifiers
     source          1..6788
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     215..237
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     234..250
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     234..250
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    270..393
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(434..739)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(536..555)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1081..1740)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1655..1674
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(1794..1800,1801..1818,1819..1824))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(1849..1972)
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        2035..2053
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2035..2052
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2240..2259)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2241..2259)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      complement(2424..3012)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2504..2523)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3335..4126)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     terminator      complement(6700..6746)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 cgatttaatt aatatgcatc aattgattta aatactagtg ttaactattt aattggggac
       61 cctagaggtc ccctttttta ttttaaaaat tttttcnaag gggacctcta gggtccccag
      121 ctggctagcg agtcatgagg tnaccatgtt ctttcctgcg ttatcccctg attctgtgga
      181 taaccgtatt accgcctttg agtgagctga taccagcgga taacaatttc acacaggaaa
      241 cagctatgac catgattacg ccaagctatc aactttgtat agaaaagttg aacgagaaac
      301 gtaaaatgat ataaatatca atatattaaa ttagattttg cataaaaaac agactacata
      361 atactgtaaa acacaacata tccagtcact atggtcgacc tgcagactgg ctgtgtataa
      421 gggagcctga catttatatt ccccagaaca tcaggttaat ggcgtttttg atgtcatttt
      481 cgcggtggct gagatcagcc acttcttccc cgataacgga gaccggcaca ctggccatat
      541 cggtggtcat catgcgccag ctttcatccc cgatatgcac caccgggtaa agttcacggg
      601 ggactttatc tgacagcaga cgtgcactgg ccagggggat caccatccgt cgcccgggcg
      661 tgtcaataat atcactctgt acatccacaa acagacgata acggctctct cttttatagg
      721 tgtaaacctt aaactgcatt tcaccagccc ctgttctcgt cggcaaaaga gccgttcatt
      781 tcaataaacc gggcgacctc agccatccct tcctgatttt ccgctttcca gcgttcggca
      841 cgcagacgac gggcttcatt ctgcatggtt gtgcttaccg aaccggagat attgacatca
      901 tatatgcctt gagcaactga tagctgtcgc tgtcaactgt cactgtaata cgctgcttca
      961 tagcatacct ctttttgaca tacttcgggt atacatatca gtatatattc ttataccgca
     1021 aaaatcagcg cgcaaatacg catactgtta tctggctttt agtaagccgg atcctctaga
     1081 ttacgccccg ccctgccact catcgcagta ctgttgtaat tcattaagca ttctgccgac
     1141 atggaagcca tcacaaacgg catgatgaac ctgaatcgcc agcggcatca gcaccttgtc
     1201 gccttgcgta taatatttgc ccatggtgaa aacgggggcg aagaagttgt ccatattggc
     1261 cacgtttaaa tcaaaactgg tgaaactcac ccagggattg gctgagacga aaaacatatt
     1321 ctcaataaac cctttaggga aataggccag gttttcaccg taacacgcca catcttgcga
     1381 atatatgtgt agaaactgcc ggaaatcgtc gtggtattca ctccagagcg atgaaaacgt
     1441 ttcagtttgc tcatggaaaa cggtgtaaca agggtgaaca ctatcccata tcaccagctc
     1501 accgtctttc attgccatac ggaattccgg atgagcattc atcaggcggg caagaatgtg
     1561 aataaaggcc ggataaaact tgtgcttatt tttctttacg gtctttaaaa aggccgtaat
     1621 atccagctga acggtctggt tataggtaca ttgagcaact gactgaaatg cctcaaaatg
     1681 ttctttacga tgccattggg atatatcaac ggtggtatat ccagtgattt ttttctccat
     1741 tttagcttcc ttagctcctg aaaatctcga cggatcctaa ctcaaaatcc acacattata
     1801 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gcggccgcca tagtgactgg
     1861 atatgttgtg ttttacagta ttatgtagtc tgttttttat gcaaaatcta atttaatata
     1921 ttgatattta tatcatttta cgtttctcgt tcaactttat tatacatagt tgcttaagct
     1981 gcaggggacc ctagaggtcc ccttttttat tttaaaaatt ttttataacc ggtgatttag
     2041 gtgacactat agaagtggaa ttcgatggcc tagctggcat cggtacgtgc cgcgcaacca
     2101 tgtagtagtc ctggagcgtg tccatctggt gttcaagctt tcgccgagca gtgcacgtac
     2161 tggacgtgtg acctgagccg cgaatacatc gacatcaacg cctgaaggcc ggcatgccat
     2221 atggaaaccg ttgtggtctc cctatagtga gtcgtattaa tttacgcgtt gcgctcggtc
     2281 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     2341 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     2401 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2461 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2521 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2581 tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg taggtatctc
     2641 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     2701 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     2761 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     2821 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     2881 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     2941 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     3001 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaactag
     3061 cctaaaggtc atccaccgga tcagcttagt aaagccctcg ctagatttta atgcggatgt
     3121 tgcgattact tcgccaacta ttgcgataac aagaaaaagc cagcctttca tgatatatct
     3181 cccaatttgt gtagggctta ttatgcacgc ttaaaaataa taaaagcaga cttgacctga
     3241 tagtttggct gtgagcaatt atgtgcttag tgcatctaac gcttgagtta agccgcgccg
     3301 cgaagcggcg tcggcttgaa cgaattgtta gacattattt gccgactacc ttggtgatct
     3361 cgcctttcac gtagtggaca aattcttcca actgatctgc gcgcgaggcc aagcgatctt
     3421 cttcttgtcc aagataagcc tgtctagctt caagtatgac gggctgatac tgggccggca
     3481 ggcgctccat tgcccagtcg gcagcgacat ccttcggcgc gattttgccg gttactgcgc
     3541 tgtaccaaat gcgggacaac gtaagcacta catttcgctc atcgccagcc cagtcgggcg
     3601 gcgagttcca tagcgttaag gtttcattta gcgcctcaaa tagatcctgt tcaggaaccg
     3661 gatcaaagag ttcctccgcc gctggaccta ccaaggcaac gctatgttct cttgcttttg
     3721 tcagcaagat agccagatca atgtcgatcg tggctggctc gaagatacct gcaagaatgt
     3781 cattgcgctg ccattctcca aattgcagtt cgcgcttagc tggataacgc cacggaatga
     3841 tgtcgtcgtg cacaacaatg gtgacttcta cagcgcggag aatctcgctc tctccagggg
     3901 aagccgaagt ttccaaaagg tcgttgatca aagctcgccg cgttgtttca tcaagcctta
     3961 cggtcaccgt aaccagcaaa tcaatatcac tgtgtggctt caggccgcca tccactgcgg
     4021 agccgtacaa atgtacggcc agcaacgtcg gttcgagatg gcgctcgatg acgccaacta
     4081 cctctgatag ttgagtcgat acttcggcga tcaccgcttc cctcatgatg tttaactttg
     4141 ttttagggcg actgccctgc tgcgtaacat cgttgctgct ccataacatc aaacatcgac
     4201 ccacggcgta acgcgcttgc tgcttggatg cccgaggcat agactgtacc ccaaaaaaac
     4261 agtcataaca agccatgaaa accgccactg cgccgttacc accgctgcgt tcggtcaagg
     4321 ttctggacca gttgcgtgag cgcatacgct acttgcatta cagcttacga accgaacagg
     4381 cttatgtcca ctgggttcgt gccttcatcc gtttccacgg tgtgcgtcac ccggcaacct
     4441 tgggcagcag cgaagtcgag gcatttctgt cctggctggc gaacgagcgc aaggtttcgg
     4501 tctccacgca tcgtcaggca ttggcggcct tgctgttctt ctacggcaag gtgctgtgca
     4561 cggatctgcc ctggcttcag gagatcggaa gacctcggcc gtcgcggcgc ttgccggtgg
     4621 tgctgacccc ggatgaagtg gttcgcatcc tcggttttct ggaaggcgag catcgtttgt
     4681 tcgcccagct tctgtatgga acgggcatgc ggatcagtga gggtttgcaa ctgcgggtca
     4741 aggatctgga tttcgatcac ggcacgatca tcgtgcggga gggcaagggc tccaaggatc
     4801 gggccttgat gttacccgag agcttggcac ccagcctgcg cgagcagggg aattgatccg
     4861 gtggatgacc gctagctggc gggaacggca atgctcctgg cggcggcggg gctggtgccc
     4921 aggtctccgc acagagcggc ggtgctggcg ctcgcggcca ggcgtggttc ttcgcgtact
     4981 gacaagaaac ccccctcttt aggactcagt gtccttggga ggggggcttt ttgcgtttca
     5041 ggaggtcttg gccagcttgg acatcgcctc agcgatagcc tcgtcgcggg cctcagacgc
     5101 catctggtac ttcatcgcca tcctaggagt cgtgtgaccg agacgggcca tcagctcctt
     5161 ggtcgtcgca cctgcctgag cggcgaacgt agcgccgaca gcgcggaggt cgtggatgcg
     5221 gagttccggc cgaccgatct tggcgtagcc acgcttcagc gacttggtga acgcggactt
     5281 cgacagccgg ttgccctgcg tcgtggtcac caggaatgcc tcggggccct tgttcatctt
     5341 cgtacggtcc ttcatgtgcg ctcggatcat ctccgcgacg tgaggcggaa ccgtcacagg
     5401 acgcttcgac cggacggtct tggcgttgcc aacgacgatc ttgttcccca cgcgggaagc
     5461 gccacggcgc acccggagct tcatcgtcat gccgtcgtcc acgatgtcct tgcggcgaag
     5521 ctcgatcagc tctccgaacc ggaggctcgt ccacgccagg atgtatgccg cgatccggta
     5581 gtgctcgaag atctcagcgg cgacgatgtc cagctcctca ggcgtcagcg cctctacgtc
     5641 gcgctcatcg gctgccttct gctcgatccg gcacgggttc tctgcgatca gcttgtcctc
     5701 gaccgctgtg ttcatcaccg cccggaggac gttgtaggca tgccggcggg cagtcgggtg
     5761 cttcctaccc atcccggccc accacgcacg caccagagct ggcgtcatct ctgtgaccgc
     5821 cacttcacct agcaccgggt agatgcggcg ctccgcgtgc ccgctgtaca gatccctggt
     5881 gccgtctgcg aggtcgcgct ccacgagcca cttccgggtg tactcctcca gcgtgatggc
     5941 gctggcggct gccttcttcg cccggtcctg tggaggggtc caggtctcca tctcgatgag
     6001 ccgcttctcg cccgcgagcc aggcttcggc gtccatcttg ttgtcgtagg tctgcagcgc
     6061 gtagtacctc acaccgtcct gcgggttgac gtatgaggct tggatcctcc cgctgcgctg
     6121 agtcttcagc gatccccatc cgcgacgtgc caactaggtc tcctctcgtc gtgaacaagg
     6181 ctaccgggtt gcaactcctg tgcaactctc aggcttcaac gcgcttctac gacctgcaat
     6241 ttctttccac ttagaggatg cagccgagag ggggtaaaaa cctatcttga ccggcccata
     6301 tatgtggtcg gcagacaccc attcttccaa actagctacg cgggttcgat tcccgtcgcc
     6361 cgctccgctg gtcagagggt gttttcgccc tctggccatt tttctttcca ggggtctgca
     6421 actcttgtgc gactcttctg acctgggcat acgcggttgc aacgcatccc tgatctggct
     6481 actttcgatg ctgacaaacg aatagagccc cccgcctgcg cgaacagacg aggggcattc
     6541 acaccagatt ggagctggtg cagtgaagag aatagaccgg gacaaggttg caccgggagt
     6601 tgcagcggtc ggaaccctcg ccgtcggcgg gctggcgttc gccctgtcgt tcacggctct
     6661 agctgatcac cgcggccatg atggccggac aaacaacaga taaaacgaaa ggcccagtct
     6721 ttcgactgag cctttcgttt tatttgatgc ctggcagtcg atcgtacgct agttaactac
     6781 gtcgacat
//
