LOCUS       sequence-414337-        5937 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP-TonB expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5937)
  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 5937)
  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: pJS65
FEATURES             Location/Qualifiers
     source          1..5937
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(88..107)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     298..319
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     rep_origin      567..1155
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1056..1075
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1309..1326
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1492..1514
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1652..1671
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(1851..2729)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        2756..3040
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     2963..2982
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             3072..3779
                     /codon_start=1
                     /product="photostable monomeric superfolder GFP (Valbuena 
                     et al., 2020)"
                     /label=msGFP2
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITGGSGGS"
     primer_bind     complement(3114..3135)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     3722..3743
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4550..4567)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      4720..4806
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4898..4925
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4944..5035
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5036..5104,5105..5896)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5254..5273)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
       61 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
      121 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
      181 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
      241 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc gggctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgttta caatttaaaa ggatctaggt gaagatcctt
      481 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
      541 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
      601 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
      661 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
      721 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
      781 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
      841 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
      901 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
      961 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     1021 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     1081 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     1141 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     1201 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     1261 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     1321 agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     1381 tcacaccgca tagggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
     1441 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     1501 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagaag gagatggcgc
     1561 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     1621 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     1681 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atctgctcat
     1741 gtttgacagc ttatcatcga tgcataatgt gcctgtcaaa tggacgaagc agggattctg
     1801 caaaccctat gctactccgt caagccgtca attgtctgat tcgttaccaa ttatgacaac
     1861 ttgacggcta catcattcac tttttcttca caaccggcac ggaactcgct cgggctggcc
     1921 ccggtgcatt ttttaaatac ccgcgagaaa tagagttgat cgtcaaaacc aacattgcga
     1981 ccgacggtgg cgataggcat ccgggtggtg ctcaaaagca gcttcgcctg gctgatacgt
     2041 tggtcctcgc gccagcttaa gacgctaatc cctaactgct ggcggaaaag atgtgacaga
     2101 cgcgacggcg acaagcaaac atgctgtgcg acgctggcga tatcaaaatt gctgtctgcc
     2161 aggtgatcgc tgatgtactg acaagcctcg cgtacccgat tatccatcgg tggatggagc
     2221 gactcgttaa tcgcttccat gcgccgcagt aacaattgct caagcagatt tatcgccagc
     2281 agctccgaat agcgcccttc cccttgcccg gcgttaatga tttgcccaaa caggtcgctg
     2341 aaatgcggct ggtgcgcttc atccgggcga aagaaccccg tattggcaaa tattgacggc
     2401 cagttaagcc attcatgcca gtaggcgcgc ggacgaaagt aaacccactg gtgataccat
     2461 tcgcgagcct ccggatgacg accgtagtga tgaatctctc ctggcgggaa cagcaaaata
     2521 tcacccggtc ggcaaacaaa ttctcgtccc tgatttttca ccaccccctg accgcgaatg
     2581 gtgagattga gaatataacc tttcattccc agcggtcggt cgataaaaaa atcgagataa
     2641 ccgttggcct caatcggcgt taaacccgcc accagatggg cattaaacga gtatcccggc
     2701 agcaggggat cattttgcgc ttcagccata cttttcatac tcccgccatt cagagaagaa
     2761 accaattgtc catattgcat cagacattgc cgtcactgcg tcttttactg gctcttctcg
     2821 ctaaccaaac cggtaacccc gcttattaaa agcattctgt aacaaagcgg gaccaaagcc
     2881 atgacaaaaa cgcgtaacaa aagtgtctat aatcacggca gaaaagtcca cattgattat
     2941 ttgcacggcg tcacactttg ctatgccata gcatttttat ccataagatt agcggatcct
     3001 acctgacgct ttttatcgca actctctact gtttctccat acccgttttt ttgggctagc
     3061 aggaggaatt catggatagc actgagagcc tgttcaccgg ggtggtgccc atcctggtcg
     3121 agctggacgg cgacgtaaac ggccacaagt tcagcgtgcg cggcgagggc gagggcgatg
     3181 ccaccaacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     3241 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     3301 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     3361 ccatcacctt caaggacgac ggcacctaca agacccgcgc cgaggtgaag ttcgagggcg
     3421 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     3481 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcacc gccgacaagc
     3541 agaagaacgg catcaaggcc aacttcaaga tccgccacaa cgtggaggac ggcagcgtgc
     3601 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     3661 acaaccacta cctgagcacc cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc
     3721 acatggtcct gctggagttc gtgaccgccg ccgggatcac tggaggcagt ggcggttctg
     3781 gtggtggtgg ttctaccctt gatttacctc gccgcttccc ctggccgacg ttactttcgg
     3841 tctgcattca tggtgctgtt gtggcgggtc tgctctatac ctcggtacat caggttattg
     3901 aactacctgc gcctgcgcag ccgatttctg tcacgatggt tacgcctgct gatctcgaac
     3961 cgccacaagc cgttcagccg ccaccggagc cggtggtaga gccagaaccg gaacctgagc
     4021 cgatccccga accgccaaaa gaagcaccgg tggtcattga aaagccgaag ccgaaaccta
     4081 agccaaaacc gaagccggtg aaaaaggtac aggagcagcc aaaacgcgat gtcaaacccg
     4141 tagagtcgcg tccggcatca ccgtttgaaa atacggcacc ggcacgcctg acatcaagta
     4201 cagcaacggc tgcaaccagc aagccggtta ccagtgtggc ttcaggacca cgcgcattaa
     4261 gccgtaatca gccgcagtat ccggcacgag cacaggcatt gcgcattgaa gggcaggtta
     4321 aagttaaatt tgacgttacg ccggatggtc gcgtggataa cgtacaaatc ctctcagcca
     4381 agcctgcgaa catgtttgag cgtgaggtga aaaatgcgat gcgcagatgg cgttatgagc
     4441 cgggtaagcc aggcagtggg attgtggtga atatcctgtt taaaattaac ggcaccaccg
     4501 aaattcagta aaagcttggc tgttttggcg gatgagagaa gattttcagc ctgatacaga
     4561 ttaaatcaga acgcagaagc ggtctgataa aacagaattt gcctggcggc agtagcgcgg
     4621 tggtcccacc tgaccccatg ccgaactcag aagtgaaacg ccgtagcgcc gatggtagtg
     4681 tggggtctcc ccatgcgaga gtagggaact gccaggcatc aaataaaacg aaaggctcag
     4741 tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg
     4801 acaaatccgc cgggagcgga tttgaacgtt gcgaagcaac ggcccggagg gtggcgggca
     4861 ggacgcccgc cataaactgc caggcatcaa attaagcaga aggccatcct gacggatggc
     4921 ctttttgcgt ttctacaaac tcttttgttt atttttctaa atacattcaa atatgtatcc
     4981 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     5041 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     5101 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     5161 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     5221 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt
     5281 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     5341 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     5401 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     5461 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     5521 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     5581 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     5641 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     5701 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     5761 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     5821 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     5881 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgattt
//
