LOCUS       sequence-334819-        4975 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 4975)
  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 4975)
  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-50-0-50%GC-Left-half-120bp
FEATURES             Location/Qualifiers
     source          1..4975
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      830..1210
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(917..936)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1127..1148
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1573..1589)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1573..1589)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1586..1608)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1597..1613
                     /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)."
     promoter        complement(join(1621..1627,1628..1645,1646..1651))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1666..1687
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1804..1821)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1975..2563)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2055..2074)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2701..3075)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     regulatory      3072..3081
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(3281..4075)
                     /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             complement(3281..4075)
                     /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(3392..3411)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4002..4021
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(4424..4726)
                     /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     complement(4526..4545)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     4849..4871
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4863..4880
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4864..4880
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4887..4906
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4887..4905
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 tcaataatcg gctgtctttc cttatcattc cttctaaatc ctcaaatgta ttatctattg
       61 acggctctaa tctattagtt gttagtgctc ctaaagatat tttagataac cttcctcaat
      121 tcctttcaac tgttgatttg ccaactgacc agatattgat tgagggtttg atatttgagg
      181 ttcagcaagg tgatgcttta gatttttcat ttgctgctgg ctctcagcgt ggcactgttg
      241 caggcggtgt taatactgac cgcctcacct ctgttttatc ttctgctggt ggttcgttcg
      301 gtatttttaa tggcgatgtt ttagggctat cagttcgcgc attaaagact aatagccatt
      361 caaaaatatt gtctgtgcca cgtattctta cgctttcagg tcagaagggt tctatctctg
      421 ttggccagaa tgtccctttt attactggtc gtgtgactgg tgaatctgcc aatgtaaata
      481 atccatttca gacgattgag cgtcaaaatg taggtatttc catgagcgtt tttcctgttg
      541 caatggctgg cggtaatatt gttctggata ttaccagcaa ggccgatagt ttgagttctt
      601 ctactcaggc aagtgatgtt attactaatc aaagaagtat tgctacaacg gttaatttgc
      661 gtgatggaca gactctttta ctcggtggcc tcactgatta taaaaacact tctcaggatt
      721 ctggcgtacc gttcctgtct aaaatccctt taatcggcct cctgtttagc tcccgctctg
      781 attctaacga ggaaagcacg ttatacgtgc tcgtcaaagc aaccatagta cgcgccctgt
      841 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
      901 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
      961 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     1021 cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga
     1081 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     1141 caaactggaa caacactcaa ccctatctcg ggctattctt ttgatttata agggattttg
     1201 ccgatttcgg aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc
     1261 gcttgctgca actctctcag cgggcacggc caggctcccg ccctcagcat ttactctggt
     1321 cacaggtatg cagctacagt caagcagtga agtaataaat tatattataa attatattat
     1381 aaattatatt atgtctagat cgactgacgt acgagtcaac gtctggatcc atttttatgg
     1441 atcgatgaag ggcgaattct gcagatatcc atcacactgg cggccgttac tagtggatcc
     1501 gagctcggta ccaagcttga tgcatagctt gagtattcta acgcgtcacc taaatagctt
     1561 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     1621 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     1681 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     1741 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     1801 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     1861 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     1921 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     1981 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     2041 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     2101 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     2161 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     2221 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     2281 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     2341 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     2401 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     2461 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     2521 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     2581 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     2641 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttagcacgtg
     2701 tcagtcctgc tcctcggcca cgaagtgcac gcagttgccg gccgggtcgc gcagggcgaa
     2761 ctcccgcccc cacggctgct cgccgatctc ggtcatggcc ggcccggagg cgtcccggaa
     2821 gttcgtggac acgacctccg accactcggc gtacagctcg tccaggccgc gcacccacac
     2881 ccaggccagg gtgttgtccg gcaccacctg gtcctggacc gcgctgatga acagggtcac
     2941 gtcgtcccgg accacaccgg cgaagtcgtc ctccacgaag tcccgggaga acccgagccg
     3001 gtcggtccag aactcgaccg ctccggcgac gtcgcgcgcg gtgagcaccg gaacggcact
     3061 ggtcaacttg gccatggtgg ccctcctcac gtgctattat tgaagcattt atcagggtta
     3121 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     3181 gcgcacattt ccccgaaaag tgccacctga tgcggtgtga aataccgcac agatgcgtaa
     3241 ggagaaaata ccgcatcagg aaattgtaag cgttaataat tcagaagaac tcgtcaagaa
     3301 ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc acgaggaagc
     3361 ggtcagccca ttcgccgcca agctcttcag caatatcacg ggtagccaac gctatgtcct
     3421 gatagcggtc cgccacaccc agccggccac agtcgatgaa tccagaaaag cggccatttt
     3481 ccaccatgat attcggcaag caggcatcgc catgggtcac gacgagatcc tcgccgtcgg
     3541 gcatgctcgc cttgagcctg gcgaacagtt cggctggcgc gagcccctga tgctcttcgt
     3601 ccagatcatc ctgatcgaca agaccggctt ccatccgagt acgtgctcgc tcgatgcgat
     3661 gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag cgtatgcagc cgccgcattg
     3721 catcagccat gatggatact ttctcggcag gagcaaggtg agatgacagg agatcctgcc
     3781 ccggcacttc gcccaatagc agccagtccc ttcccgcttc agtgacaacg tcgagcacag
     3841 ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg cgctgcctcg tcttgcagtt
     3901 cattcagggc accggacagg tcggtcttga caaaaagaac cgggcgcccc tgcgctgaca
     3961 gccggaacac ggcggcatca gagcagccga ttgtctgttg tgcccagtca tagccgaata
     4021 gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc atcttgttca atcatgcgaa
     4081 acgatcctca tcctgtctct tgatcagagc ttgatcccct gcgccatcag atccttggcg
     4141 gcaagaaagc catccagttt actttgcagg gcttcccaac cttaccagag ggcgccccag
     4201 ctggcaattc cggttcgctt gctgtccata aaaccgccca gtctagctat cgccatgtaa
     4261 gcccactgca agctacctgc tttctctttg cgcttgcgtt ttcccttgtc cagatagccc
     4321 agtagctgac attcatccgg ggtcagcacc gtttctgcgg actggctttc tacgtgaaaa
     4381 ggatctaggt gaagatcctt tttgataatc tcatgcctga catttatatt ccccagaaca
     4441 tcaggttaat ggcgtttttg atgtcatttt cgcggtggct gagatcagcc acttcttccc
     4501 cgataacgga gaccggcaca ctggccatat cggtggtcat catgcgccag ctttcatccc
     4561 cgatatgcac caccgggtaa agttcacggg agactttatc tgacagcaga cgtgcactgg
     4621 ccagggggat caccatccgt cgccccggcg tgtcaataat atcactctgt acatccacaa
     4681 acagacgata acggctctct cttttatagg tgtaaacctt aaactgccgt acgtataggc
     4741 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     4801 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
     4861 gttgtaaaac gacggccagt gaattgtaat acgactcact atagggcgaa ttgggccctc
     4921 tagatgcatg ctcgagcggc cgccagtgtg atggatatct gcagaattcg ccctt
//
