LOCUS       sequence-416353-        6687 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TolQR expression vector for purification (TolQ-TolR-TEV-Strep_His).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6687)
  AUTHORS   Williams-Jones DP, Webby MN, Press CE, Gradon JM, Armstrong SR, 
            Szczepaniak J, Kleanthous C
  TITLE     Tunable force transduction through the Escherichia coli cell 
            envelope.
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Nov 21;120(47):e2306707120. doi: 
            10.1073/pnas.2306707120. Epub 2023 Nov 16.
  PUBMED    37972066
REFERENCE   2  (bases 1 to 6687)
  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: "2023-11-21"; volume: "120(47):e2306707120. 
            doi"; pages: " 10.1073/pnas.2306707120. Epub 2023 Nov 16"
COMMENT     Sequence Label: pCEP13
FEATURES             Location/Qualifiers
     source          1..6687
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        164..282
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     CDS             1434..1454
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1467..1490
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1527..1550
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1551..1568
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1625..1642)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1795..1881
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1973..2000
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             complement(2072..2887)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2072..2887)
                     /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
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2795..2814
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2879..2898
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(4947..5795)
                     /codon_start=1
                     /gene="E. coli rhaR"
                     /product="transcriptional activator of rhaR and rhaS"
                     /label=rhaR
                     /note="needed for rhamnose-dependent activation of the rhaB
                     promoter"
                     /translation="MAHQLKLLKDDFFASDQQAVAVADRYPQDVFAEHTHDFCELVIVW
                     RGNGLHVLNDRPYRITRGDLFYIHADDKHSYASVNDLVLQNIIYCPERLKLNLDWQGAI
                     PGFNASAGQPHWRLGSMGMAQARQVIGQLEHESSQHVPFANEMAELLFGQLVMLLNRHR
                     YTSDSLPPTSSETLLDKLITRLAASLKSPFALDKFCDEASCSERVLRQQFRQQTGMTIN
                     QYLRQVRVCHAQYLLQHSRLLISDISTECGFEDSNYFSVVFTRETGMTPLAGRHLNSQK
                     D"
     CDS             complement(join(5869..6687,1..18))
                     /codon_start=1
                     /gene="E. coli rhaS"
                     /product="positive regulator of the rhaB promoter
                     "
                     /label=rhaS
                     /note="activated by L-rhamnose"
                     /translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
                     GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLNQLLPQ
                     ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
                     ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
                     LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
ORIGIN
        1 actatgtaat acggtcatac tggcctcctg atgtcgtcaa cacggcgaaa tagtaatcac
       61 gaggtcaggt tcttacctta aattttcgac ggaaaaccac gtaaaaaacg tcgatttttc
      121 aagatacagc gtgaattttc aggaaatgcg gtgagcatca catcaccaca attcagcaaa
      181 ttgtgaacat catcacgttc atctttccct ggttgccaat ggcccatttt cttgtcagta
      241 acgagaaggt cgcgaatcca ggcgcttttt agactggtcg taatgaaatt caggaggaat
      301 ccatggtgac tgacatgaat atccttgatt tgttcctgaa ggctagcctt ctggttaaac
      361 ttatcatgtt gattttgatt ggtttttcaa tcgcatcttg ggccattatt atccagcgga
      421 cccgtattct taacgcagcg gcgcgcgaag ccgaagcgtt tgaagataaa ttctggtctg
      481 gaatcgaact ctctcgcctc tatcaagaga gccaggggaa acgggataat ctgactggtt
      541 cggaacaaat cttttacagc gggttcaaag agtttgtgcg cctgcatcgt gccaatagcc
      601 atgcgccgga agccgtagtg gaaggggcgt cgcgtgctat gcgtatctcc atgaaccgtg
      661 aacttgaaaa tctggaaacg cacattccgt tcctcggtac ggttggctcc atcagcccgt
      721 atattggtct gtttggtacg gtctggggga tcatgcacgc ctttatcgcc ctcggggcgg
      781 taaaacaagc aacactgcaa atggttgcgc ccggtatcgc agaagcgttg attgcgactg
      841 caattggtct gtttgccgct atcccggcag ttatggccta caaccgcctc aaccagcgcg
      901 taaacaaact ggaactgaat tacgacaact ttatggaaga gtttaccgcg attctgcacc
      961 gccaggcgtt taccgttagc gagagcaaca aggggtaagc catggccaga gcgcgtggac
     1021 gaggtcgtcg cgatctcaag tccgaaatca acattgtacc gttgctggac gtactgctgg
     1081 tgctgttgct gatctttatg gcgacagcgc ccatcatcac ccagagcgtg gaggtcgatc
     1141 tgccagacgc tactgaatca caggcggtga gcagtaacga taatccgcca gtgattgttg
     1201 aagtgtctgg tattggtcag tacaccgtgg tggttgagaa agatcgcctg gagcgtttac
     1261 caccagagca ggtggtggcg gaagtgtcca gccgtttcaa ggccaacccg aaaacggtct
     1321 ttctgatcgg tggcgcaaaa gatgtgcctt acgatgaaat aattaaagca ctgaacttgt
     1381 tacatagtgc gggtgtgaaa tcggttggtt taatgacgca gcctatcctc gaggaaaacc
     1441 tgtacttcca gggtcaattc gggagctgga gccacccgca gttcgagaaa ggtggaggtt
     1501 ccggaggtgg atcgggaggt ggatcgtgga gccacccgca gttcgaaaaa catcaccatc
     1561 accatcacta aaacagaaaa accgactagc ttggctgttt tggcggatga gagaagattt
     1621 tcagcctgat acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg
     1681 gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta
     1741 gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata
     1801 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     1861 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
     1921 ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc
     1981 atcctgacgg atggcctttt tgcgtttcta caaactcttt tgtttatttt tctaaataca
     2041 ttcaaatatg tatccgctca tgagactagg cttagaaaaa ctcatcgagc atcaaatgaa
     2101 actgcaattt attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta
     2161 atgaaggaga aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg
     2221 cgattccgac tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt
     2281 tatcaagtga gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat
     2341 gcatttcttt ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg
     2401 catcaaccaa accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc
     2461 tgttaaaagg acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg
     2521 catcaacaat attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc
     2581 cggggatcgc agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg
     2641 tcggaagagg cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat
     2701 tggcgacgct acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca
     2761 atcgatagat tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata
     2821 aatcagcatc catgttggaa tttaatcgcg gcctagagca agacgtttcc cgttgaatat
     2881 ggctcataac accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgacc
     2941 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     3001 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     3061 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3121 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     3181 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3241 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     3301 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacggcgcc tgtagtgcca
     3361 tttaccccca ttcactgcca gagccgtgag cgcagcgaac tgaatgtcac gaaaaagaca
     3421 gcgactcagg tgcctgatgg tcggagacaa aaggaatatt cagcgatttg cccgagcttg
     3481 cgagggtgct acttaagcct ttagggtttt aaggtctgtt ttgtagagga gcaaacagcg
     3541 tttgcgacat ccttttgtaa tactgcggaa ctgactaaag tagtgagtta tacacagggc
     3601 tgggatctat tctttttatc tttttttatt ctttctttat tctataaatt ataaccactt
     3661 gaatataaac aaaaaaaaca cacaaaggtc tagcggaatt tacagagggt ctagcagaat
     3721 ttacaagttt tccagcaaag gtctagcaga atttacagat acccacaact caaaggaaaa
     3781 ggactagctc actcggtcgc tacgctccgg gcgtgagact gcggcgggcg ctgcggacac
     3841 atacaaagtt acccacagat tccgtggata agcaggggac taacatgtga ggcaaaacag
     3901 cagggccgcg ccggtggcgt ttttccatag gctccgccct cctgccagag ttcacataaa
     3961 cagacgcttt tccggtgcat ctgtgggagc cgtgaggctc aaccatgaat ctgacagtac
     4021 gggcgaaacc cgacaggact taaagatccc caccgtttcc ggcgggtcgc tccctcttgc
     4081 gctctcctgt tccgaccctg ccgtttaccg gatacctgtt ccgcctttct cccttacggg
     4141 aagtgtggcg ctttctcata gctcacacac tggtatctcg gctcggtgta ggtcgttcgc
     4201 tccaagctgg gctgtaagca agaactcccc gttcagcccg actgctgcgc cttatccgat
     4261 aactgttcac ttgagtccaa cccggaaaag cacggtaaaa cgccactggc agcagccatt
     4321 ggtaactggg agttcgcaga ggatttgttt agctaaacac gcggttgctc ttgaagtgtg
     4381 cgccaaagtc cggctacact ggaaggacag atttggttgc tgtgctctgc gaaagccagt
     4441 taccacggtt aagcagttcc ccaactgact taaccttcga tcaaaccacc tccccaggtg
     4501 gttttttcga aggttaagtc agttggggaa ctgcttaacc ttcgatcaaa ccacctcccc
     4561 aggtggtttt ttcgtttaca gggcaaaaga ttacgcgcag aaaaaaagga tctcaagaag
     4621 atcctttgat cttttctact gaaccgctct agctttcagt gcaatttatc tcttcaaatg
     4681 tagcacctga agtcagcccc atacgatata agttgtaatt ctcatgttag tcatgccccg
     4741 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     4801 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4861 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4921 gcgtattggg cgcccgataa ggttaattaa tctttctgcg aattgagatg acgcccagcc
     4981 agtggcgtca tcccggtttc ccgggtaaac accaccgaaa aatagttact atcttcaaag
     5041 ccacattcgg tcgaaatatc actgattaac aggcggctat gctggagaag atattgcgca
     5101 tgacacactc tgacctgtcg cagatattga ttgatggtca ttccagtctg ctggcgaaat
     5161 tgctgacgca aaacgcgctc actgcacgat gcctcatcac aaaatttatc cagcgcaaag
     5221 ggacttttca ggctagccgc cagccgggta atcagcttat ccagcaacgt ttcgctggat
     5281 gttggcggca acgaatcact ggtgtaacga tggcgattca gcaacatcac caactgcccg
     5341 aacagcaact cagccatttc gttagcaaac ggcacatgct gactactttc atgctcaagc
     5401 tgaccaataa cctgccgcgc ctgcgccatc cccatgctac ctaagcgcca gtgtggttgc
     5461 cctgcgctgg cgttaaatcc cggaatcgcc ccctgccagt caagattcag cttcagacgc
     5521 tccgggcaat aaataatatt ctgcaaaacc agatcgttaa cggaagcgta ggagtgttta
     5581 tcatcagcat gaatgtaaaa gagatcgcca cgggtaatgc gataagggcg atcgttgagt
     5641 acatgcaggc cattaccgcg ccagacaatc accagctcac aaaaatcatg tgtatgttca
     5701 gcaaagacat cttgcggata acggtcagcc acagcgactg cctgctggtc gctggcaaaa
     5761 aaatcatctt tgagaagttt taactgatgc gccaccgtgg ctacctcggc cagagaacga
     5821 agttgattat tcgcaatatg gcgtacaaat acgttgagaa gattcgcgtt attgcagaaa
     5881 gccatcccgt ccctggcgaa tatcacgcgg tgaccagtta aactctcggc gaaaaagcgt
     5941 cgaaaagtgg ttactgtcgc tgaatccaca gcgataggcg atgtcagtaa cgctggcctc
     6001 gctgtggcgt agcagatgtc gggctttcat cagtcgcagg cggttcaggt atcgctgagg
     6061 cgtcagtccc gtttgctgct taagctgccg atgtagcgta cgcagtgaaa gagaaaattg
     6121 atccgccacg gcatcccaat tcacctcatc ggcaaaatgg tcctccagcc aggccagaag
     6181 caagttgaga cgtgatgcgc tgttttccag gttctcctgc aaactgcttt tacgcagcaa
     6241 gagcagtaat tgcataaaca agatctcgcg actggcggtc gagggtaaat cattttcccc
     6301 ttcctgctgt tccatctgtg caaccagctg tcgcacctgc tgcaatacgc tgtggttaac
     6361 gcgccagtga gacggatact gcccatccag ctcttgtggc agcaactgat tcagcccggc
     6421 gagaaactga aatcgatccg gcgagcgata cagcacattg gtcagacaca gattatcggt
     6481 atgttcatac agatgccgat catgatcgcg tacgaaacag accgtgccac cggtgatggt
     6541 atagggctgc ccattaaaca catgaatacc cgtgccatgt tcgacaatca caatttcatg
     6601 aaaatcatga tgatgttcag gaaaatccgc ctgcgggagc cggggttcta tcgccacgga
     6661 cgcgttacca gacggaaaaa aatccac
//
