LOCUS       sequence-471782-        3023 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression vector for RoTq-Off.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3023)
  AUTHORS   Rosen PC, Glaser A, Martinez-Francois JR, Lim DC, Brooks DJ, Fu P, 
            Kim E, Kern D, Yellen G
  TITLE     Mechanism and application of thiol-disulfide redox biosensors with a
            fluorescence-lifetime readout.
  JOURNAL   Proc Natl Acad Sci U S A. 2025 May 13;122(19):e2503978122. doi: 
            10.1073/pnas.2503978122. Epub 2025 May 6.
  PUBMED    40327692
REFERENCE   2  (bases 1 to 3023)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2025-05-13"; volume: "122(19):e2503978122. 
            doi"; pages: " 10.1073/pnas.2503978122. Epub 2025 May 6"
COMMENT     Sequence Label: pRsetB-RoTq-Off
FEATURES             Location/Qualifiers
     source          1..3023
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     138..155
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     240..259
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        240..258
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             290..312
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             326..346
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     primer_bind     1165..1186
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1349..1453
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1454..2248
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             1454..2248
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1508..1527)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2118..2137
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      2419..3007
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2908..2927
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
       61 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
      121 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca
      181 aaccgcctct ccccgcgcgt tggccgattc attaatgcag gatctcgatc ccgcgaaatt
      241 aatacgactc actataggga gaccacaacg gtttccctct agaaataatt ttgtttaact
      301 ttaagaagga gatatacata tgaaacatca ccatcatcat catcatggtg gagctagcat
      361 ggtgagcaaa ggcgaagaac tgtttaccgg cgtggtgccg attctggtgg aactggatgg
      421 cgatgtgaac ggccataaat ttagcgtgag cggcgaaggc gaaggcgatg cgacctatgg
      481 caaactgacc ctgaaattta ttagcaccac cggcaaactg ccggtgccgt ggccgaccct
      541 ggtgaccacc ctgtcctggg gcgtgcagtg ctttgcgcgc tatccggatc atatgaaacg
      601 ccatgatttt tttaaaagcg cgatgccgga aggctatgtg caggaacgca ccattttttt
      661 taaagatgat ggcaactata aaacccgcgc ggaagtgaaa tttgaaggcg ataccctggt
      721 gaaccgcatt gaactgaaag gcattgattt taaagaagat ggcaacattc tgggccataa
      781 actggaatat aacggtgggt gcgaagataa cgtgtatatt accgcggata aacagaaaaa
      841 cggcattaaa gcgaacttta aaattcgcca taacattgaa gatggcggcg tgcagctggc
      901 ggatcattat cagcagaaca ccccgattgg cgatggcccg gtgctgctgc cggataacca
      961 ttatctgagc acctgcagca aactgagcaa agatccgaac gaaaaacgcg atcatatggt
     1021 gtttctggaa tttgtgaccg cggcgggcat taccctgggc atggatgaac tgtataaata
     1081 actcgagggg cccaagctgt agtgggccat cgccctgata gacggttttt cgccctttga
     1141 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     1201 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     1261 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     1321 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     1381 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     1441 aaaaaggaag agtatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     1501 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     1561 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     1621 cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     1681 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     1741 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     1801 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     1861 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     1921 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     1981 gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     2041 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     2101 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     2161 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     2221 ctatcgcctt cttgacgagt tcttctgact gtcagaccaa gtttactcat atatacttta
     2281 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     2341 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     2401 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2461 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2521 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     2581 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     2641 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     2701 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     2761 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     2821 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     2881 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     2941 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     3001 atggaaaaac gccagcaacg cgg
//
