LOCUS       sequence-95199-e        5222 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PTac K1FR sigma fragment expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5222)
  AUTHORS   Segall-Shapiro TH, Meyer AJ, Ellington AD, Sontag ED, Voigt CA
  TITLE     A 'resource allocator' for transcription based on a highly 
            fragmented T7 RNA polymerase.
  JOURNAL   Mol Syst Biol. 2014 Jul 30;10(7):742.
  PUBMED    25080493
REFERENCE   2  (bases 1 to 5222)
  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: "Mol Syst 
            Biol."; date: "2014-07-30"; volume: "10(7)"; pages: "742"
COMMENT     Sequence Label: pTHSSd_25
FEATURES             Location/Qualifiers
     source          1..5222
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      1040..1071
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     rep_origin      1582..2126
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     2027..2046
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2243..2260
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             2835..3626
                     /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(3753..3782)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     CDS             complement(3845..4927)
                     /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     4889..4908
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5062..5067,5068..5083,5084..5090)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    5096..5115
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     misc_RNA        5131..5180
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
ORIGIN
        1 atgagcatcg cggcgaccct ggagaacgat ctggcgcgtc tggaaaacga aaacgctcgt
       61 ctcgaaaaag acatcgcgaa cctggaacgt gacctggcga aactggagcg tgaagaagcg
      121 tacttcggag gttcaggtgg taagaacact ggtgaaatct ctgagaaagt caagctgggc
      181 actaaggcac tggctggtca atggctggct tacggtgtta ctcgcagtgt gactaagcgt
      241 tcagtcatga cgctggctta cgggtccaaa gagttcggct tccgtcaaca agtgctggaa
      301 gataccattc agccagctat tgattccggc aagggtctga tgttcactca gccgaatcag
      361 gctgctggat acatggctaa gctgatttgg gaatctgtga gcgtgacggt ggtagctgcg
      421 gttgaagcaa tgaactggct taagtctgct gctaagctgc tggctgctga ggtcaaagat
      481 aagaagactg gagagattct tcgcaagcgt tgcgctgtgc attgggtaac tcctgatggt
      541 ttccctgtgt ggcaggaata caagaagcct attcagacgc gcttgaacct gaggttcctc
      601 ggttcgttca acctccagcc gaccgtcaac accaacaaag atagcgagat tgatgcacac
      661 aaacaggagt ctggtatcgc tcctaacttt gtacacagcc aagacggtag ccaccttcgt
      721 aagactgtag tgtgggcaca cgagaagtac ggaatcgaat cttttgcact gattcacgac
      781 tccttcggta cgattccggc tgacgctgcg aacctgttca aagcagtgcg cgaaactatg
      841 gttgacacat atgagtcttg tgatgtactg gctgatttct acgaccagtt cgctgaccag
      901 ttgcacgagt ctcaattgga caaaatgcca gcacttccgg ctaaaggtaa cttgaacctc
      961 cgtgacatct tagagtcgga cttcgcgttc gcgtaacgct gcaggagtca ctaagggtta
     1021 gttagttaga ttagcagaaa gtcaaaagcc tccgaccgga ggcttttgac taaaacttcc
     1081 cttggggtta tcattggggc tcactcaaag gcggtaatca gataaaaaaa atccttagct
     1141 ttcgctaagg atgatttctg ctagagatgg aatagactgg atggaggcgg ataaagttgc
     1201 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     1261 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     1321 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     1381 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     1441 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     1501 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     1561 ccccttaata agatgatctt cttgagatcg ttttggtctg cgcgtaatct cttgctctga
     1621 aaacgaaaaa accgccttgc agggcggttt ttcgaaggtt ctctgagcta ccaactcttt
     1681 gaaccgaggt aactggcttg gaggagcgca gtcaccaaaa cttgtccttt cagtttagcc
     1741 ttaaccggcg catgacttca agactaactc ctctaaatca attaccagtg gctgctgcca
     1801 gtggtgcttt tgcatgtctt tccgggttgg actcaagacg atagttaccg gataaggcgc
     1861 agcggtcgga ctgaacgggg ggttcgtgca tacagtccag cttggagcga actgcctacc
     1921 cggaactgag tgtcaggcgt ggaatgagac aaacgcggcc ataacagcgg aatgacaccg
     1981 gtaaaccgaa aggcaggaac aggagagcgc acgagggagc cgccagggga aacgcctggt
     2041 atctttatag tcctgtcggg tttcgccacc actgatttga gcgtcagatt tcgtgatgct
     2101 tgtcaggggg gcggagccta tggaaaaacg gctttgccgc ggccctctca cttccctgtt
     2161 aagtatcttc ctggcatctt ccaggaaatc tccgccccgt tcgtaagcca tttccgctcg
     2221 ccgcagtcga acgaccgagc gtagcgagtc agtgagcgag gaagcggaat atatcctgta
     2281 tcacatattc tgctgacgca ccggtgcagc cttttttctc ctgccacatg aagcacttca
     2341 ctgacaccct catcagtgcc aacatagtaa gccagtatac actccgctag cgctgaggtc
     2401 tgcctcgtga agaaggtgtt gctgactcat accaggcctg aatcgcccca tcatccagcc
     2461 agaaagtgag ggagccacgg ttgatgagag ctttgttgta ggtggaccag ttggtgattt
     2521 tgaacttttg ctttgccacg gaacggtctg cgttgtcggg aagatgcgtg atctgatcct
     2581 tcaactcagc aaaagttcga tttattcaac aaagccacgt tgtgtctcaa aatctctgat
     2641 gttacattgc acaagataaa aatatatcat catgaacaat aaaactgtct gcttacataa
     2701 acagtaatac aaggggtgtt tactagaggt tgatcgggca cgtaagaggt tccaactttc
     2761 accataatga aataagatca ctaccgggcg tattttttga gttatcgaga ttttcaggag
     2821 ctaaggaagc taaaatgagg gaagcggtga tcgccgaagt atcgactcaa ctatcagagg
     2881 tagttggcgt catcgagcgc catctcgaac cgacgttgct ggccgtacat ttgtacggct
     2941 ccgcagtgga tggcggcctg aagccacaca gtgatattga tttgctggtt acggtgaccg
     3001 taaggcttga tgaaacaacg cggcgagctt tgatcaacga ccttttggaa acttcggctt
     3061 cccctggaga gagcgagatt ctccgcgctg tagaagtcac cattgttgtg cacgacgaca
     3121 tcattccgtg gcgttatcca gctaagcgcg aactgcaatt tggagaatgg cagcgcaatg
     3181 acattcttgc aggtatcttc gagccagcca cgatcgacat tgatctggct atcttgctga
     3241 caaaagcaag agaacatagc gttgccttgg taggtccagc ggcggaggaa ctctttgatc
     3301 cggttcctga acaggatcta tttgaggcgc taaatgaaac cttaacgcta tggaactcgc
     3361 cgcccgactg ggctggcgat gagcgaaatg tagtgcttac gttgtcccgc atttggtaca
     3421 gcgcagtaac cggcaaaatc gcgccgaagg atgtcgctgc cgactgggca atggagcgcc
     3481 tgccggccca gtatcagccc gtcatacttg aagctagaca ggcttatctt ggacaagaag
     3541 aagatcgctt ggcctcgcgc gcagatcagt tggaagaatt tgtccactac gtgaaaggcg
     3601 agatcaccaa ggtagtcggc aaataatact agctccggca aaaaaacggg caaggtgtca
     3661 ccaccctgcc ctttttcttt aaaaccgaaa agattacttc gcgtttgcca cctgacgtct
     3721 aagaaaagga atattcagca atttgcccgt gccgaagaaa ggcccacccg tgaaggtgag
     3781 ccagtgagtt gattgctacg taatgtcggc caattcgcgc taacttacat taattgcgtt
     3841 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     3901 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca
     3961 gtgagactgg caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc
     4021 ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga
     4081 tataacatga gctatcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc
     4141 gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca
     4201 gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca
     4261 tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt
     4321 tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg
     4381 cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcctcat
     4441 gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa
     4501 cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga
     4561 tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc
     4621 cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa
     4681 tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca
     4741 gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg
     4801 ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca
     4861 cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg
     4921 gtttcatatt caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa
     4981 aggttttgcg ccattcgatg gcgcgccgct tcgtcaggcc acatagcttt cttgttctga
     5041 tcggaacgat cgttggctgt gttgacaatt aatcatcggc tcgtataatg tgtggaattg
     5101 tgagcgctca caattctatg gactatgttt ctgtcaccgg atgtgctttc cggtctgatg
     5161 agtccgtgag gacgaaacag cctctacaaa taattttgtt taatcacaca ggaaagtact
     5221 ag
//
