LOCUS       sequence-450047-        6125 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the HUH endonuclease WDV, used as a no ligand control for 
            DNA tension sensor experiments.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6125)
  AUTHORS   Pawlak MR, Smiley AT, Ramirez MP, Kelly MD, Shamsan GA, Anderson SM,
            Smeester BA, Largaespada DA, Odde DJ, Gordon WR
  TITLE     RAD-TGTs: high-throughput measurement of cellular mechanotype via 
            rupture and delivery of DNA tension probes.
  JOURNAL   Nat Commun. 2023 Apr 28;14(1):2468. doi: 10.1038/s41467-023-38157-6.
  PUBMED    37117218
REFERENCE   2  (bases 1 to 6125)
  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: "Nat 
            Commun."; date: "2023-04-28"; volume: "14(1):2468. doi"; pages: " 
            10.1038/s41467-023-38157-6"
COMMENT     Sequence Label: WDV
FEATURES             Location/Qualifiers
     source          1..6125
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(114..1196)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1158..1177
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1197..1274)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1480..1499
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1583..1602
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1583..1601
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1602..1626
                     /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)."
     RBS             1641..1663
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1682..1699
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1712..2002
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="SDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLR
                     RLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             2439..2456
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2509..2527)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2523..2570
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2607..3062
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2694..2713)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2904..2925
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3088..3192
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3193..3261,3262..4053)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3411..3430)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4224..4812
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4713..4732
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4966..4983
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4998..5137
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5063..5082)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5223..5245
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5239..5430)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg
       61 gtcgagatcc cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc
      121 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
      181 ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc
      241 aacagctgat tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg
      301 gtttgcccca gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag
      361 ctgtcttcgg tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac
      421 tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg
      481 ggaacgatgc cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag
      541 tcgccttccc gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca
      601 gccagacgca gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg
      661 tgacccaatg cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata
      721 atactgttga tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag
      781 gcagcttcca cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg
      841 acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct
      901 accatcgaca ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca
      961 atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt
     1021 ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct
     1081 tccacttttt cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg
     1141 gtctgataag agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc
     1201 accaccctga attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc
     1261 cattcgatgg tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca
     1321 gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga
     1381 gatggcgccc aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc
     1441 gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc
     1501 gccagcaacc gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat
     1561 cgagatctcg atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac
     1621 aattcccctc tagaaataat tttgtttaac tttaagaagg agatatacca tgcgtggttc
     1681 tcaccaccac caccaccata tggctagcgg atcggactca gaagtcaatc aagaagctaa
     1741 gccagaggtc aagccagaag tcaagcctga gactcacatc aatttaaagg tgtccgatgg
     1801 atcttcagag atcttcttca agatcaaaaa gaccactcct ttaagaaggc tgatggaagc
     1861 gttcgctaaa agacagggta aggaaatgga ctccttaaga ttcttgtacg acggtattag
     1921 aattcaagct gatcagaccc ctgaagattt ggacatggag gataacgata ttattgaggc
     1981 tcacagagaa cagattggtg gatccatggc aagcagcagc acaccgcgtt ttcgtgttta
     2041 tagcaaatac ctgtttctga cctatccgca gtgtaccctg gaaccgcagt atgcactgga
     2101 tagcctgcgt accctgctga acaaatatga accgctgtat attgcagcag ttcgtgaact
     2161 gcatgaagat ggtagtccgc atctgcatgt tctggttcag aataaactgc gtgcaagcat
     2221 taccaatccg aatgcactga atctgcgtat ggataccagt ccgtttagca tttttcatcc
     2281 gaatattcag gcagccaaag attgtaatca ggtgcgtgat tatatcacca aagaagttga
     2341 tagcgacgtt aataccgcag aatggggcac ctttgttgca gttagcacac cgggtcgtaa
     2401 agatcgtgat gcagatctcg agtaataagc ttgtcgagca ccaccaccac caccactgag
     2461 atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat
     2521 aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag
     2581 gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg
     2641 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2701 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2761 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2821 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2881 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2941 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     3001 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
     3061 ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     3121 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     3181 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     3241 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     3301 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     3361 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     3421 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     3481 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     3541 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     3601 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     3661 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     3721 gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     3781 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     3841 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     3901 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     3961 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     4021 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     4081 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     4141 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     4201 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     4261 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     4321 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     4381 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     4441 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     4501 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     4561 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     4621 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     4681 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     4741 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     4801 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     4861 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     4921 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     4981 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     5041 caccgcaatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtata
     5101 cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc
     5161 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     5221 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagct
     5281 gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct gttcatccgc
     5341 gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa agcgggccat
     5401 gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt
     5461 catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg ttactgatga
     5521 tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg
     5581 ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag atgtaggtgt
     5641 tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg tgcagggcgc
     5701 tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc atgttgttgc
     5761 tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc
     5821 attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg acaggagcac
     5881 gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg
     5941 tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc
     6001 aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca
     6061 gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc
     6121 gacga
//
