LOCUS       sequence-334821-        6915 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Force-activating hairpin for 3'-to-5' helicase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6915)
  AUTHORS   Taylor AMK, Okoniewski SR, Uyetake L, Perkins TT
  TITLE     Force-Activated DNA Substrates for In Situ Generation of ssDNA and 
            Designed ssDNA/dsDNA Structures in an Optical-Trapping Assay.
  JOURNAL   Methods Mol Biol. 2022;2478:273-312. doi: 
            10.1007/978-1-0716-2229-2_10.
  PUBMED    36063324
REFERENCE   2  (bases 1 to 6915)
  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: "Methods Mol
            Biol."; date: "2022"; volume: "2478:273-312. doi"; pages: " 
            10.1007/978-1-0716-2229-2_10"
COMMENT     Sequence Label: hp-75%GC-40bp
FEATURES             Location/Qualifiers
     source          1..6915
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      834..1214
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(921..940)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1131..1152
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             1485..2675
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     complement(1837..1856)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(2993..3012)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2994..3012)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3019..3036)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3019..3035)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3028..3050)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             3173..3475
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3354..3373
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             3824..4618
                     /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             3824..4618
                     /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(3878..3897)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4488..4507
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             complement(join(4868..5659,5660..5728))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5491..5510
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5852..6440
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6341..6360
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6594..6611
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6728..6749
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6764..6769,6770..6787,6788..6794)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6802..6818
                     /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)."
     primer_bind     6807..6829
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6826..6842
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6826..6842
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        6863..6881
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 catgtcaata atcggctgtc tttccttatc attccttcta aatcctcaaa tgtattatct
       61 attgacggct ctaatctatt agttgttagt gctcctaaag atattttaga taaccttcct
      121 caattccttt caactgttga tttgccaact gaccagatat tgattgaggg tttgatattt
      181 gaggttcagc aaggtgatgc tttagatttt tcatttgctg ctggctctca gcgtggcact
      241 gttgcaggcg gtgttaatac tgaccgcctc acctctgttt tatcttctgc tggtggttcg
      301 ttcggtattt ttaatggcga tgttttaggg ctatcagttc gcgcattaaa gactaatagc
      361 cattcaaaaa tattgtctgt gccacgtatt cttacgcttt caggtcagaa gggttctatc
      421 tctgttggcc agaatgtccc ttttattact ggtcgtgtga ctggtgaatc tgccaatgta
      481 aataatccat ttcagacgat tgagcgtcaa aatgtaggta tttccatgag cgtttttcct
      541 gttgcaatgg ctggcggtaa tattgttctg gatattacca gcaaggccga tagtttgagt
      601 tcttctactc aggcaagtga tgttattact aatcaaagaa gtattgctac aacggttaat
      661 ttgcgtgatg gacagactct tttactcggt ggcctcactg attataaaaa cacttctcag
      721 gattctggcg taccgttcct gtctaaaatc cctttaatcg gcctcctgtt tagctcccgc
      781 tctgattcta acgaggaaag cacgttatac gtgctcgtca aagcaaccat agtacgcgcc
      841 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
      901 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
      961 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     1021 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
     1081 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     1141 gttccaaact ggaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat
     1201 tttgccgatt tcggaaccac catcaaacag gattttcgcc tgctggggca aaccagcgtg
     1261 gaccgcttgc tgcaactctc tcagcgggca cggccaggct cccgccctca gcatttgcga
     1321 ccgacggagc ctccgtggga cggagcgtgc catttttatg gcacgctccg tcccacggag
     1381 gctccgtcgg tcgctttgct gtctagagga tccgactatc gacctcagcg ctttaatgcg
     1441 gtagtttatc acagttaaat tgctaacgca gtcaggcacc gtgtatgaaa tctaacaatg
     1501 cgctcatcgt catcctcggc accgtcaccc tggatgctgt aggcataggc ttggttatgc
     1561 cggtactgcc gggcctcttg cgggatatcg tccattccga cagcatcgcc agtcactatg
     1621 gcgtgctgct agcgctatat gcgttgatgc aatttctatg cgcacccgtt ctcggagcac
     1681 tgtccgaccg ctttggccgc cgcccagtcc tgctcgcttc gctacttgga gccactatcg
     1741 actacgcgat catggcgacc acacccgtcc tgtggatcct ctacgccgga cgcatcgtgg
     1801 ccggcatcac cggcgccaca ggtgcggttg ctggcgccta tatcgccgac atcaccgatg
     1861 gggaagatcg ggctcgccac ttcgggctca tgagcgcttg tttcggcgtg ggtatggtgg
     1921 caggccccgt ggccggggga ctgttgggcg ccatctcctt gcatgcacca ttccttgcgg
     1981 cggcggtgct caacggcctc aacctactac tgggctgctt cctaatgcag gagtcgcata
     2041 agggagagcg tcgaccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg
     2101 cgcggggcat gactatcgtc gccgcactta tgactgtctt ctttatcatg caactcgtag
     2161 gacaggtgcc ggcagcgctc tgggtcattt tcggcgagga ccgctttcgc tggagcgcga
     2221 cgatgatcgg cctgtcgctt gcggtattcg gaatcttgca cgccctcgct caagccttcg
     2281 tcactggtcc cgccaccaaa cgtttcggcg agaagcaggc cattatcgcc ggcatggcgg
     2341 ccgacgcgct gggctacgtc ttgctggcgt tcgcgacgcg aggctggatg gccttcccca
     2401 ttatgattct tctcgcttcc ggcggcatcg ggatgcccgc gttgcaggcc atgctgtcca
     2461 ggcaggtaga tgacgaccat cagggacagc ttcaaggatc gctcgcggct cttaccagcc
     2521 taacttcgat cattggaccg ctgatcgtca cggcgattta tgccgcctcg gcgagcacat
     2581 ggaacgggtt ggcatggatt gtaggcgccg ccctatacct tgtctgcctc cccgcgttgc
     2641 gtcgcggtgc atggagccgg gccacctcga cctgaatgga agccggcggc acctcgctaa
     2701 cggattcacc actccaagaa ttggagccaa tcaattcttg cggagaactg tgaatgcgca
     2761 aaccaaccct tggcagaaca tatccatcgc gtccgccatc tccagcagcc gcacgcggcg
     2821 catctcgggc agcgttgggt cctggccacg ggtgcgcatg atcgtgctcc tgtcgttgag
     2881 gacccggcta ggctggcggg gttgccttac tggttagcag aatgaatcac cgatacgcga
     2941 gcgaacgtga agcgactgca agggcgaatt cgcggccgct aaattcaatt cgccctatag
     3001 tgagtcgtat tacaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     3061 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     3121 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctatacgtac ggcagtttaa
     3181 ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac agagtgatat
     3241 tattgacacg ccggggcgac ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga
     3301 taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct ggcgcatgat
     3361 gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg ctgatctcag
     3421 ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa tataaatgtc
     3481 aggcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttcacgtaga aagccagtcc
     3541 gcagaaacgg tgctgacccc ggatgaatgt cagctactgg gctatctgga caagggaaaa
     3601 cgcaagcgca aagagaaagc aggtagcttg cagtgggctt acatggcgat agctagactg
     3661 ggcggtttta tggacagcaa gcgaaccgga attgccagct ggggcgccct ctggtaaggt
     3721 tgggaagccc tgcaaagtaa actggatggc tttcttgccg ccaaggatct gatggcgcag
     3781 gggatcaagc tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg
     3841 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
     3901 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
     3961 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg
     4021 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
     4081 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
     4141 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
     4201 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
     4261 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     4321 gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt
     4381 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     4441 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     4501 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     4561 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgaat
     4621 tattaacgct tacaatttcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     4681 acaccgcatc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     4741 tctaaataca ttcaaatatg tatccgctca tgagattatc aaaaaggatc ttcacctaga
     4801 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     4861 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     4921 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     4981 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     5041 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     5101 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     5161 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     5221 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     5281 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     5341 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     5401 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     5461 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     5521 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     5581 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     5641 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     5701 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     5761 atcagggtta ttgtctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5821 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5881 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5941 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     6001 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6061 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     6121 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     6181 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     6241 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     6301 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     6361 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     6421 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     6481 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     6541 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     6601 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     6661 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     6721 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     6781 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     6841 accatgatta cgccaagctc agaattaacc ctcactaaag ggactagtcc tgcaggttta
     6901 aacgaattcg ccctt
//
