LOCUS       sequence-334822-        6921 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Force-activating hairpin for 5'-to-3' helicase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6921)
  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 6921)
  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%GC-40bp-Revbindsite
FEATURES             Location/Qualifiers
     source          1..6921
                     /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             1491..2681
                     /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(1843..1862)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(2999..3018)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3000..3018)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3025..3042)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3025..3041)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3034..3056)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             3179..3481
                     /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     3360..3379
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             3830..4624
                     /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             3830..4624
                     /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(3884..3903)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4494..4513
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             complement(join(4874..5665,5666..5734))
                     /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     5497..5516
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5858..6446
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6347..6366
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6600..6617
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6734..6755
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6770..6775,6776..6793,6794..6800)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6808..6824
                     /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     6813..6835
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6832..6848
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6832..6848
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        6869..6887
                     /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 tcacctcagc gctgtctaga ggatccgact atcgaacccg
     1321 tctagatcga ctgacgtacg agtcaacgtc tggatccatt tttatggatc cagacgttga
     1381 ctcgtacgtc agtcgatcta gactttcctc agcgcgggca cggccaggct cccgcgcttt
     1441 aatgcggtag tttatcacag ttaaattgct aacgcagtca ggcaccgtgc atgaaatcta
     1501 acaatgcgct catcgtcatc ctcggcaccg tcaccctgga tgctgtaggc ataggcttgg
     1561 ttatgccggt actgccgggc ctcttgcggg atatcgtcca ttccgacagc atcgccagtc
     1621 actatggcgt gctgctagcg ctatatgcgt tgatgcaatt tctatgcgca cccgttctcg
     1681 gagcactgtc cgaccgcttt ggccgccgcc cagtcctgct cgcttcgcta cttggagcca
     1741 ctatcgacta cgcgatcatg gcgaccacac ccgtcctgtg gatcctctac gccggacgca
     1801 tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca
     1861 ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta
     1921 tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc
     1981 ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt
     2041 cgcataaggg agagcgtcga ccgatgccct tgagagcctt caacccagtc agctccttcc
     2101 ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac
     2161 tcgtaggaca ggtgccggca gcgctctggg tcattttcgg cgaggaccgc tttcgctgga
     2221 gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc ctcgctcaag
     2281 ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt atcgccggca
     2341 tggcggccga cgcgctgggc tacgtcttgc tggcgttcgc gacgcgaggc tggatggcct
     2401 tccccattat gattcttctc gcttccggcg gcatcgggat gcccgcgttg caggccatgc
     2461 tgtccaggca ggtagatgac gaccatcagg gacagcttca aggatcgctc gcggctctta
     2521 ccagcctaac ttcgatcatt ggaccgctga tcgtcacggc gatttatgcc gcctcggcga
     2581 gcacatggaa cgggttggca tggattgtag gcgccgccct ataccttgtc tgcctccccg
     2641 cgttgcgtcg cggtgcatgg agccgggcca cctcgacctg aatggaagcc ggcggcacct
     2701 cgctaacgga ttcaccactc caagaattgg agccaatcaa ttcttgcgga gaactgtgaa
     2761 tgcgcaaacc aacccttggc agaacatatc catcgcgtcc gccatctcca gcagccgcac
     2821 gcggcgcatc tcgggcagcg ttgggtcctg gccacgggtg cgcatgatcg tgctcctgtc
     2881 gttgaggacc cggctaggct ggcggggttg ccttactggt tagcagaatg aatcaccgat
     2941 acgcgagcga acgtgaagcg actgcaaggg cgaattcgcg gccgctaaat tcaattcgcc
     3001 ctatagtgag tcgtattaca attcactggc cgtcgtttta caacgtcgtg actgggaaaa
     3061 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     3121 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctat acgtacggca
     3181 gtttaaggtt tacacctata aaagagagag ccgttatcgt ctgtttgtgg atgtacagag
     3241 tgatattatt gacacgccgg ggcgacggat ggtgatcccc ctggccagtg cacgtctgct
     3301 gtcagataaa gtctcccgtg aactttaccc ggtggtgcat atcggggatg aaagctggcg
     3361 catgatgacc accgatatgg ccagtgtgcc ggtctccgtt atcggggaag aagtggctga
     3421 tctcagccac cgcgaaaatg acatcaaaaa cgccattaac ctgatgttct ggggaatata
     3481 aatgtcaggc atgagattat caaaaaggat cttcacctag atccttttca cgtagaaagc
     3541 cagtccgcag aaacggtgct gaccccggat gaatgtcagc tactgggcta tctggacaag
     3601 ggaaaacgca agcgcaaaga gaaagcaggt agcttgcagt gggcttacat ggcgatagct
     3661 agactgggcg gttttatgga cagcaagcga accggaattg ccagctgggg cgccctctgg
     3721 taaggttggg aagccctgca aagtaaactg gatggctttc ttgccgccaa ggatctgatg
     3781 gcgcagggga tcaagctctg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     3841 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     3901 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     3961 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc
     4021 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     4081 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     4141 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     4201 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     4261 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     4321 gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct
     4381 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     4441 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     4501 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     4561 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     4621 ctgaattatt aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg
     4681 tatttcacac cgcatcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     4741 tatttttcta aatacattca aatatgtatc cgctcatgag attatcaaaa aggatcttca
     4801 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     4861 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     4921 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     4981 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     5041 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     5101 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     5161 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     5221 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     5281 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     5341 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     5401 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     5461 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     5521 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     5581 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     5641 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     5701 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     5761 gcatttatca gggttattgt ctcatgacca aaatccctta acgtgagttt tcgttccact
     5821 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     5881 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     5941 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     6001 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     6061 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     6121 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     6181 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     6241 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     6301 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     6361 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     6421 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     6481 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     6541 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     6601 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     6661 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     6721 agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta
     6781 tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca
     6841 gctatgacca tgattacgcc aagctcagaa ttaaccctca ctaaagggac tagtcctgca
     6901 ggtttaaacg aattcgccct t
//
