LOCUS       sequence-12885-e        7315 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 7315)
  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 7315)
  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: pGMCKq26-10M-hyg
FEATURES             Location/Qualifiers
     source          1..7315
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(105..151)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     427..443
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    463..483
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     537..553
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     538..554
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1492..1516
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     1515..1534
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     protein_bind    1571..1604
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    3081..3114
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3175..3191)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3199..3215
                     /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)."
     promoter        complement(join(3223..3229,3230..3247,3248..3253))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3268..3289
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    complement(3311..3335)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3353..3373)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     promoter        3436..3454
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3436..3453
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3640..3659)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3641..3659)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      complement(3742..4330)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3822..3841)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4662..5477)
                     /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
                     KPHAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHNLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5385..5404
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5469..5488
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 ggagttgcag cggtcggaac cctcgccgtc ggcgggctgg cgttcgccct gtcgttcacg
       61 gctctagctg atcaccgcgg ccatgatggc cggacaaaca acagataaaa cgaaaggccc
      121 agtctttcga ctgagccttt cgttttattt gatgcctggc agtcgatcgt acgctagtta
      181 actacgtcga catcgattta attaatatgc atcaattgat ttaaatacta gtgttaacta
      241 tttaattggg gaccctagag gtcccctttt ttattttaaa aattttttcn aaggggacct
      301 ctagggtccc cagctggcta gcgagtcatg aggtnaccat gttctttcct gcgttatccc
      361 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccagc ggataacaat
      421 ttcacacagg aaacagctat gaccatgatt acgccaagct atcaactttg tatagaaaag
      481 ttgcatgacc aacttcgata acgttctcgg ctcgatgatc cccggcgggc cccccctcga
      541 ggtcgacggt atcgatttaa ttaatatgca tcaattgatt taaatactag tgttaactat
      601 ttaattgggg accctagagg tccccttttt tattttaaaa attttttcna aggggacctc
      661 tagggtcccc agctggctag cgagtcatga ggtnaccatg gtcattagga gccgctctcg
      721 cacttcagct gcttctccaa gccgcagatg atcagctcca ggccgaacaa gaaggccggc
      781 tcggcgccct ggtggtcgaa cagctcgatg gcctggcgca gcaacggcgg catgctgtcg
      841 gtggtcgggg tctcgcgctc ctccttggcg acctggtgct cctggtcctc caacacgcag
      901 cccagggtga agtggcccac ggccgacagg gcgtacaacg cgttctccag cgagaagccc
      961 tgctggcaca ggaaggccaa ctggttctcc agggtctcgt actgcttctc ggtcggccgg
     1021 gtgcccaggt gcaccttggc gccgtcgcgg tggctcagca aggcgcagcg gaagctcttg
     1081 gcgttgttgc gcaggaagtc ctgccagctc tcgccctcca gcgggcagaa gtgggtgtgg
     1141 tggcggtcca acatctcgat ggccagggcg tcgagcaggg cccgcttgtt cttgacgtgc
     1201 cagtacaggg tcggctgctc gacgcccagc ttctgggcca gcttgcgggt ggtcaggccc
     1261 tcgatgccga cctcgttcag cagctccagg gcggagttga tgaccttgga cttgtccagg
     1321 cgggacattg cgaagtgatt cctcctgatg gtaccggatc cgtggcagga gggtcgcctg
     1381 cgtcggcaac tcgcccaggc tacacgagag ccctcgactg cgaaaatgcc gaccaggcac
     1441 caccgcactc caggcttaac ccggtgcgac gatccaatat tacgagatct gacaagtttg
     1501 tacaaaaaag caggcggcat cagagcagat tgtactgaga gtgcaccata tgggatccgg
     1561 taccgaattc ataacttcgt ataatgtatg ctatacgaag ttatatcgat gggtctgacg
     1621 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     1681 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     1741 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     1801 tatttcgttc atccatagtt gcctgactcc ccggacgtgg ccgaccagcc cgtcatcgtc
     1861 aacgcctgac cgcggtgcgg acaggccgtg tcgcgaccgg ccgtgcggaa ttaagccggc
     1921 ccgtaccctg tgaatagagg tccgctgtga cacaagaatc cctgttactt ctcgaccgta
     1981 ttgattcgga tgattcctac gcgagcctgc ggaacgacca ggagttctgg gagccgctgg
     2041 cccgccgagc cctggaggag ctcgggctgc cggtgccgcc ggtgctgcgg gtgcccggcg
     2101 agagcaccaa ccccgtactg gtcggcgagc ccgacccggt gatcaagctg ttcggcgagc
     2161 actggtgcgg tccggagagc ctcgcgtcgg agtcggaggc gtacgcggtc ctggcggacg
     2221 ccccggtgcc ggtgccccgc ctcctcggcc gcggcgagct gcggcccggc accggagcct
     2281 ggccgtggcc ctacctggtg atgagccgga tgaccggcac cacctggcgg tccgcgatgg
     2341 acggcacgac cgaccggaac gcgctgctcg ccctggcccg cgaactcggc cgggtgctcg
     2401 gccggctgca cagggtgccg ctgaccggga acaccgtgct caccccccat tccgaggtct
     2461 tcccggaact gctgcgggaa cgccgcgcgg cgaccgtcga ggaccaccgc gggtggggct
     2521 acctctcgcc ccggctgctg gaccgcctgg aggactggct gccggacgtg gacacgctgc
     2581 tggccggccg cgaaccccgg ttcgtccacg gcgacctgca cgggaccaac atcttcgtgg
     2641 acctggccgc gaccgaggtc accgggatcg tcgacttcac cgacgtctat gcgggagact
     2701 cccgctacag cctggtgcaa ctgcatctca acgccttccg gggcgaccgc gagatcctgg
     2761 ccgcgctgct cgacggggcg cagtggaagc ggaccgagga cttcgcccgc gaactgctcg
     2821 ccttcacctt cctgcacgac ttcgaggtgt tcgaggagac cccgctggat ctctccggct
     2881 tcaccgatcc ggaggaactg gcgcagttcc tctgggggcc gccggacacc gcccccggcg
     2941 cctgacgccc cgggccgccc ggcggcggac gctcagcagc ggcgggcgtg aaaggccctg
     3001 gcatcctcga tcatctcctc cagggtggtc ggccgagctt ccatcccagc tcggcaagga
     3061 tcaacgagga cagtaggcct ataacttcgt ataatgtatg ctatacgaag ttatgaattc
     3121 ggtaccggat cctctagagt cgacctgcag gcatgcaagc ttggcgtaat catggtcata
     3181 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     3241 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3301 ctcactgccc acccagcttt cttgtacaaa gtggttaatt aagcgcaagc ttcaacttta
     3361 ttatacatag ttgcttaagc tgcaggggac cctagaggtc ccctttttta ttttaaaaat
     3421 tttttataac cggtgattta ggtgacacta tagaagtgga attcgatggc ctagctggca
     3481 tcggtacgtg ccgcgcaacc atgtagtagt cctggagcgt gtccatctgg tgttcaagct
     3541 ttcttctaca acaacccgct gctgtcgtta cgccgcctcc ttggattcga cgacggacat
     3601 ggcgtagtcg tgcatgccat atggaaaccg ttgtggtctc cctatagtga gtcgtattaa
     3661 tttacgcgtt tcactggccg acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     3721 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     3781 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     3841 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     3901 cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta ggtatctcag
     3961 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     4021 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     4081 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     4141 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     4201 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     4261 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     4321 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaactagtg
     4381 aggtctgcct cgtgaagaag gtgttgctga ctcataccag gcctgaatcg ccccatcatc
     4441 cagccagaaa gtgagggagc cacggttgat gagagctttg ttgtaggtgg accagttggt
     4501 gattttgaac ttttgctttg ccacggaacg gtctgcgttg tcgggaagat gcgtgatctg
     4561 atccttcaac tcagcaaaag ttcgatttat tcaacaaagc cgccgtcccg tcaagtcagc
     4621 gtaatgctct gccagtgtta caaccaatta accaattctg attagaaaaa ctcatcgagc
     4681 atcaaatgaa actgcaattt attcatatca ggattatcaa taccatattt ttgaaaaagc
     4741 cgtttctgta atgaaggaga aaactcaccg aggcagttcc ataggatggc aagatcctgg
     4801 tatcggtctg cgattccgac tcgtccaaca tcaatacaac ctattaattt cccctcgtca
     4861 aaaataaggt tatcaagtga gaaatcacca tgagtgacga ctgaatccgg tgagaatggc
     4921 aaaagattat gcatttcttt ccagacttgt tcaacaggcc agccattacg ctcgtcatca
     4981 aaatcactcg catcaaccaa accgttattc attcgtgatt gcgcctgagc gagacgaaat
     5041 acgcgatcgc tgttaaaagg acaattacaa acaggaatcg aatgcaaccg gcgcaggaac
     5101 actgccagcg catcaacaat attttcacct gaatcaggat attcttctaa tacctggaat
     5161 gctgttttcc cggggatcgc agtggtgagt aaccatgcat catcaggagt acggataaaa
     5221 tgcttgatgg tcggaagagg cataaattcc gtcagccagt ttagtctgac catctcatct
     5281 gtaacatcat tggcaacgct acctttgcca tgtttcagaa acaactctgg cgcatggggc
     5341 ttcccataca agcgatagat tgtcgcacct gattgcccga cattatcgcg agcccattta
     5401 tacccatata aatcagcatc catgttggaa tttaatcgcg gcctcgagca agacgtttcc
     5461 cgttgaatat ggctcataac accccttgta ttactgttta tgtaagcaga cagttttatt
     5521 gttcatgatg atatattttt atcttgtgca atgtaacatc agagattttg agacacaacg
     5581 tcgctttgtt gctagctggc gggaacggca atgctcctgg cggcggcggg gctggtgccc
     5641 aggtctccgc acagagcggc ggtgctggcg ctcgcggcca ggcgtggttc ttcgcgtact
     5701 gacaagaaac ccccctcttt aggactcagt gtccttggga ggggggcttt ttgcgtttca
     5761 ggaggtcttg gccagcttgg acatcgcctc agcgatagcc tcgtcgcggg cctcagacgc
     5821 catctggtac ttcatcgcca tcctaggagt cgtgtgaccg agacgggcca tcagctcctt
     5881 ggtcgtcgca cctgcctgag cggcgaacgt agcgccgaca gcgcggaggt cgtggatgcg
     5941 gagttccggc cgaccgatct tggcgtagcc acgcttcagc gacttggtga acgcggactt
     6001 cgacagccgg ttgccctgcg tcgtggtcac caggaatgcc tcggggccct tgttcatctt
     6061 cgtacggtcc ttcatgtgcg ctcggatcat ctccgcgacg tgaggcggaa ccgtcacagg
     6121 acgcttcgac cggacggtct tggcgttgcc aacgacgatc ttgttcccca cgcgggaagc
     6181 gccacggcgc acccggagct tcatcgtcat gccgtcgtcc acgatgtcct tgcggcgaag
     6241 ctcgatcagc tctccgaacc ggaggctcgt ccacgccagg atgtatgccg cgatccggta
     6301 gtgctcgaag atctcagcgg cgacgatgtc cagctcctca ggcgtcagcg cctctacgtc
     6361 gcgctcatcg gctgccttct gctcgatccg gcacgggttc tctgcgatca gcttgtcctc
     6421 gaccgctgtg ttcatcaccg cccggaggac gttgtaggca tgccggcggg cagtcgggtg
     6481 cttcctaccc atcccggccc accacgcacg caccagagct ggcgtcatct ctgtgaccgc
     6541 cacttcacct agcaccgggt agatgcggcg ctccgcgtgc ccgctgtaca gatccctggt
     6601 gccgtctgcg aggtcgcgct ccacgagcca cttccgggtg tactcctcca gcgtgatggc
     6661 gctggcggct gccttcttcg cccggtcctg tggaggggtc caggtctcca tctcgatgag
     6721 ccgcttctcg cccgcgagcc aggcttcggc gtccatcttg ttgtcgtagg tctgcagcgc
     6781 gtagtacctc acaccgtcct gcgggttgac gtatgaggct tggatcctcc cgctgcgctg
     6841 agtcttcagc gatccccatc cgcgacgtgc caactaggtc tcctctcgtc gtgaacaagg
     6901 ctaccgggtt gcaactcctg tgcaactctc aggcttcaac gcgcttctac gacctgcaat
     6961 ttctttccac ttagaggatg cagccgagag ggggtaaaaa cctatcttga ccggcccata
     7021 tatgtggtcg gcagacaccc attcttccaa actagctacg cgggttcgat tcccgtcgcc
     7081 cgctccgctg gtcagagggt gttttcgccc tctggccatt tttctttcca ggggtctgca
     7141 actcttgtgc gactcttctg acctgggcat acgcggttgc aacgcatccc tgatctggct
     7201 actttcgatg ctgacaaacg aatagagccc cccgcctgcg cgaacagacg aggggcattc
     7261 acaccagatt ggagctggtg cagtgaagag aatagaccgg gacaaggttg caccg
//
