LOCUS       sequence-361604-        4055 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SpySwitch in the bacterial cytoplasm. SpySwitch binds 
            SpyTag-, SpyTag002- or SpyTag003-fusions non-covalently for affinity
            purification, allowing gentle pH or temperature release..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4055)
  AUTHORS   Vester SK, Rahikainen R, Khairil Anuar INA, Hills RA, Tan TK, 
            Howarth M
  TITLE     SpySwitch enables pH- or heat-responsive capture and release for 
            plug-and-display nanoassembly.
  JOURNAL   Nat Commun. 2022 Jun 28;13(1):3714. doi: 10.1038/s41467-022-31193-8.
  PUBMED    35764623
REFERENCE   2  (bases 1 to 4055)
  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: "2022-06-28"; volume: "13(1):3714. doi"; pages: " 
            10.1038/s41467-022-31193-8"
COMMENT     Sequence Label: SpySwitch
FEATURES             Location/Qualifiers
     source          1..4055
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             41..63
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             546..563
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(616..634)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      630..677
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      714..1169
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(801..820)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1011..1032
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1195..1299
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1300..1368,1369..2160)
                     /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(1518..1537)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2331..2919
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2820..2839
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3073..3090
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3105..3244
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3170..3189)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3330..3352
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3346..3537)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     join(4046..4055,1..10)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(4046..4055,1..9)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 cactataggg agaccacaac ggtttccctc tagaaataat tttgtttaac tttaagaagg
       61 agatatacat atgtcgtact accatcacca tcaccatcac ggtagcggcg gtagcggcgc
      121 catggtaacc accttatcag gtttatcagg tgagcaaggt ccgtccggtg atatgacaac
      181 tgaagaagat agtgctaccc atattcactt ccataaacat gatgaggacg gccgtgagtt
      241 agctggtgca actatggagt tgcgtgattg ctctggtaaa actattagta catggattac
      301 agatggacat gtgaaggatt tctacctgta tccaggaaaa tatacatttg tcgcgaccgc
      361 agcaccagac ggtcatcacg tagcaactcc aattgaattt acaattaatg aggacggtca
      421 ggttactgta gatggtgaag caactgaagg tgaccctcat actggatcca gtggtagcta
      481 ataaatggtt gatgcttgag gatccgaatt cgagctccgt cgacaagctt gcggccgcac
      541 tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg
      601 agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
      661 tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat gggacgcgcc
      721 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
      781 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
      841 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
      901 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
      961 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     1021 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     1081 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     1141 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
     1201 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     1261 ccctggtaaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     1321 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     1381 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     1441 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     1501 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     1561 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     1621 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     1681 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     1741 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     1801 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg
     1861 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     1921 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     1981 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     2041 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     2101 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     2161 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     2221 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     2281 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     2341 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     2401 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     2461 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     2521 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2581 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2641 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2701 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2761 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2821 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2881 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2941 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     3001 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     3061 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     3121 ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct
     3181 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     3241 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     3301 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     3361 gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga
     3421 ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agcttctcca gaagcgttaa
     3481 tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga
     3541 tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag
     3601 gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg
     3661 taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca
     3721 gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca
     3781 gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg
     3841 gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct
     3901 tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc
     3961 tagccgggtc ctcaacgaca ggagcacaat catgcgcacc cgtggccagg acccaacgct
     4021 gcccgagatc tcgatcccgc gaaattaata cgact
//
