LOCUS       sequence-270569-        7478 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Sigma 70 S366C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7478)
  AUTHORS   Mumm JP, Friedman LJ, Gelles J
  TITLE     Mechanism of upstream promoter element stimulation of transcription 
            at a ribosomal RNA promoter determined by single-molecule imaging
  JOURNAL   bioRxiv 2020
REFERENCE   2  (bases 1 to 7478)
  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: "bioRxiv 
            2020"
COMMENT     Sequence Label: rpoDs366C
FEATURES             Location/Qualifiers
     source          1..7478
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             100..117
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             127..144
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2271..2289)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2357..2461
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2462..2530,2531..3322)
                     /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     complement(2680..2699)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3493..4081
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3982..4001
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4235..4252
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4267..4406
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4332..4351)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4492..4514
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4508..4699)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     7054..7073
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7093..7170)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7376..7395
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg ggcagcagcc atcatcatca tcatcacagc
      121 agcggcctgg tgccgcgcgg cagccatatg gagcaaaacc cgcagtcaca gctgaaactt
      181 cttgtcaccc gtggtaagga gcaaggctat ctgacctatg ccgaggtcaa tgaccatctg
      241 ccggaagata tcgtcgattc agatcagatc gaagacatca tccaaatgat caacgacatg
      301 ggcattcagg tgatggaaga agcaccggat gccgatgatc tgatgctggc tgaaaacacc
      361 gcggacgaag atgctgccga agccgccgcg caggtgcttt ccagcgtgga atctgaaatc
      421 gggcgcacga ctgacccggt acgcatgtac atgcgtgaaa tgggcaccgt tgaactgttg
      481 acccgcgaag gcgaaattga catcgctaag cgtattgaag acgggatcaa ccaggttcaa
      541 tcctccgttg ctgaatatcc ggaagcgatc acctatctgc tggaacagta cgatcgtgtt
      601 gaagcagaag aagcgcgtct gtccgatctg atcaccggct ttgttgaccc gaacgcagaa
      661 gaagatctgg cacctaccgc cactcacgtc ggttctgagc tttcccagga agatctggac
      721 gatgacgaag atgaagacga agaagatggc gatgacgaca gcgccgatga tgacaacagc
      781 atcgacccgg aactggctcg cgaaaaattt gcggaactac gcgctcagta cgttgtaacg
      841 cgtgacacca tcaaagcgaa aggtcgcagt cacgctaccg ctcaggaaga gatcctgaaa
      901 ctgtctgaag tattcaaaca gttccgcctg gtgccgaagc agtttgacta cctggtcaac
      961 agcatgcgcg tcatgatgga ccgcgttcgt acgcaagaac gtctgatcat gaagctctcc
     1021 gttgagcagt ccaaaatgcc gaagaaaaac ttcattaccc tgtttaccgg caacgaaacc
     1081 agcgatacct ggttcaacgc ggcaattgcg atgaacaagc cgtggtcgga aaaactgcac
     1141 gatgtctctg aagaagtgca tcgcgccctg caaaaactgc agcagattga agaagaaacc
     1201 ggcctgacca tcgagcaggt taaagatatc aaccgtcgta tgtgcatcgg tgaagcgaaa
     1261 gcccgccgtg cgaagaaaga gatggttgaa gcgaacttac gtctggttat ttctatcgct
     1321 aagaaataca ccaaccgtgg cttgcagttc cttgacctga ttcaggaagg caacatcggt
     1381 ctgatgaaag cggttgataa attcgaatac cgccgtggtt acaagttctc cacctacgca
     1441 acctggtgga tccgtcaggc gatcacccgc tctatcgcgg atcaggcgcg caccatccgt
     1501 attccggtgc atatgattga gaccatcaac aagctcaacc gtatttctcg ccagatgctg
     1561 caagagatgg gccgtgaacc gacgccggaa gaactggctg aacgtatgct gatgccggaa
     1621 gacaagatcc gcaaagtgct gaagatcgcc aaagagccaa tctccatgga aacgccgatc
     1681 ggtgatgatg aagattcgca tctgggggat ttcatcgagg ataccaccct cgagctgccg
     1741 ctggattctg cgaccaccga aagcctgcgt gcggcaacgc acgacgtgct ggctggcctg
     1801 accgcgcgtg aagcaaaagt tctgcgtatg cgtttcggta tcgatatgaa caccgactac
     1861 acgctggaag aagtgggtaa acagttcgac gttacccgcg aacgtatccg tcagatcgaa
     1921 gcgaaggcgc tgcgcaaact gcgtcacccg agccgttctg aagtgctgcg tagcttcctg
     1981 gacgattaat cggtaggccg gatcaggcgt tacgccgcac ccggcactag gccctctgca
     2041 caaacgccac cttttcggtg gcgtttttta tcgcccacgc actaccagcg cctggtccag
     2101 ctcgcgatac gcttcaacca gtttctccag tgaaacgcga cttaaaccgc tgggatttgg
     2161 cagcacccaa atctgcgtcg aaccaatggt gagcgtttgt ttcccccact gtgcagccca
     2221 agcttatcga tgataagctg tcaaacatga gaattcttga agacgaaagg gcctcgtgat
     2281 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     2341 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     2401 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     2461 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     2521 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     2581 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     2641 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     2701 ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     2761 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     2821 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     2881 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     2941 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     3001 gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     3061 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     3121 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     3181 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     3241 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     3301 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     3361 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     3421 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     3481 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     3541 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     3601 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     3661 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     3721 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     3781 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     3841 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     3901 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     3961 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     4021 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     4081 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     4141 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca ttgagtgagc
     4201 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     4261 agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcaatgg
     4321 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat
     4381 cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
     4441 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     4501 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     4561 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     4621 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     4681 ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa
     4741 tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc
     4801 ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa
     4861 aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta
     4921 gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg
     4981 tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag
     5041 acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac
     5101 cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca
     5161 cccgtggcca ggacccaacg ctgcccgaga tgcgccgcgt gcggctgctg gagatggcgg
     5221 acgcgatgga tatgttctgc caagggttgg tttgcgcatt cacagttctc cgcaagaatt
     5281 gattggctcc aattcttgga gtggtgaatc cgttagcgag gtgccgccgg cttccattca
     5341 ggtcgaggtg gcccggctcc atgcaccgcg acgcaacgcg gggaggcaga caaggtatag
     5401 ggcggcgcct acaatccatg ccaacccgtt ccatgtgctc gccgaggcgg cataaatcgc
     5461 cgtgacgatc agcggtccaa tgatcgaagt taggctggta agagccgcga gcgatccttg
     5521 aagctgtccc tgatggtcgt catctacctg cctggacagc atggcctgca acgcgggcat
     5581 cccgatgccg ccggaagcga gaagaatcat aatggggaag gccatccagc ctcgcgtcgc
     5641 gaacgccagc aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa tggcctgctt
     5701 ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat
     5761 tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc
     5821 gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt
     5881 cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa
     5941 ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat
     6001 tgcgttgcgc tcactgcccg ctttccagtc gggaaactgt cgtgccagct gcattaatga
     6061 atcggccaac gcgcggggag aggcggtttg cgtattgggc gccagggtgg tttttctttt
     6121 caccagtgag acgggcaaca gctgattgcc cttcaccgcc tggccctgag agagttgcag
     6181 caagcggtcc acgctggttt gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg
     6241 cgggatataa catgagctgt cttcggtatc gtcgtatccc actaccgaga tatccgcacc
     6301 aacgcgcagc ccggactcgg taatggcgcg cattgcgccc agcgccatct gatcgttggc
     6361 aaccagcatc gcagtgggaa cgatgccctc attcagcatt tgcatggttt gttgaaaacc
     6421 ggacatggca ctccagtcgc cttcccgttc cgctatcggc tgaatttgat tgcgagtgag
     6481 atatttatgc cagccagcca gacgcagacg cgccgagaca gaacttaatg ggcccgctaa
     6541 cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc
     6601 ttcatgggag aaaataatac tgttgatggg tgtctggtca gagacatcaa gaaataacgc
     6661 cggaacatta gtgcaggcag cttccacagc aatggcatcc tggtcatcca gcggatagtt
     6721 aatgatcagc ccactgacgc gttgcgcgag aagattgtgc accgccgctt tacaggcttc
     6781 gacgccgctt cgttctacca tcgacaccac cacgctggca cccagttgat cggcgcgaga
     6841 tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc agactggagg tggcaacgcc
     6901 aatcagcaac gactgtttgc ccgccagttg ttgtgccacg cggttgggaa tgtaattcag
     6961 ctccgccatc gccgcttcca ctttttcccg cgttttcgca gaaacgtggc tggcctggtt
     7021 caccacgcgg gaaacggtct gataagagac accggcatac tctgcgacat cgtataacgt
     7081 tactggtttc acattcacca ccctgaattg actctcttcc gggcgctatc atgccatacc
     7141 gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg acgctctccc ttatgcgact
     7201 cctgcattag gaagcagccc agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga
     7261 atggtgcatg caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac
     7321 ccacgccgaa acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga
     7381 tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc
     7441 gtccggcgta gaggatcgag atctcgatcc cgcgaaat
//
