LOCUS       sequence-205343-        3636 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Monomeric SpyTag-6×-His alpha-hemolysin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3636)
  AUTHORS   Stranges PB, Palla M, Kalachikov S, Nivala J, Dorwart M, Trans A, 
            Kumar S, Porel M, Chien M, Tao C, Morozova I, Li Z, Shi S, Aberra A,
            Arnold C, Yang A, Aguirre A, Harada ET, Korenblum D, Pollard J, Bhat
            A, Gremyachinskiy D, Bibillo A, Chen R, Davis R, Russo JJ, Fuller 
            CW, Roever S, Ju J, Church GM
  TITLE     Design and characterization of a nanopore-coupled polymerase for 
            single-molecule DNA sequencing by synthesis on an electrode array.
  JOURNAL   Proc Natl Acad Sci U S A. 2016 Nov 1;113(44):E6749-E6756. doi: 
            10.1073/pnas.1608271113. Epub 2016 Oct 11.
  PUBMED    27729524
REFERENCE   2  (bases 1 to 3636)
  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"; doi: 
            "10.1073/pnas.1608271113"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2016-11-1- 1"; volume: "113"; issue: "44"; pages: 
            "E6749-E6756"
COMMENT     Sequence Label: pEXP5-aHL_spytag_his
FEATURES             Location/Qualifiers
     source          1..3636
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             52..74
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             994..1032
                     /codon_start=1
                     /product="peptide derived from the CnaB2 adhesin domain of 
                     Streptococcus pyogenes FbaB"
                     /label=SpyTag
                     /note="forms a covalent isopeptide bond with the SpyCatcher
                     protein (Zakeri et al., 2012)"
                     /translation="AHIVMVDAYKPTK"
     CDS             1039..1056
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1106..1153
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1198..1216)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1284..1388
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1389..1457,1458..2249)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1607..1626)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2420..3008
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2909..2928
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3162..3179
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3259..3278)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 taatacgact cactataggg agacacacga cggtttccct ctagaaataa ttttgtttaa
       61 ctttaagaag gagataccct tatggcagat tctgatatta atattaaaac cggtactaca
      121 gatattggaa gcaatactac agtaaaaaca ggtgatttag tcacttatga taaagaaaat
      181 ggcatgcaca aaaaagtatt ttatagtttt atcgatgata aaaatcacaa taaaaaactg
      241 ctagttatta gaacgaaagg taccattgct ggtcaatata gagtttatag cgaagaaggt
      301 gctaacaaaa gtggtttagc ctggccttca gcctttaagg tacagttgca actacctgat
      361 aatgaagtag ctcaaatatc tgattactat ccaagaaatt cgattgatac aaaagagtat
      421 atgagtactt taacttatgg attcaacggt aatgttactg gtgatgatac aggaaaaatt
      481 ggcggcctta ttggtgcaaa tgtttcgatt ggtcatacac tgaaatatgt tcaacctgat
      541 ttcaaaacaa ttttagagag cccaactgat aaaaaagtag gctggaaagt gatatttaac
      601 aatatggtga atcaaaattg gggaccatat gatagagatt cttggaaccc ggtatatggc
      661 aatcaacttt tcatgaaaac tagaaatggt tctatgaaag cagcagagaa cttccttgat
      721 cctaacaaag caagttctct attatcttca gggttttcac cagacttcgc tacagttatt
      781 actatggata gaaaagcatc caaacaacaa acaaatatag atgtaatata cgaacgagtt
      841 cgtgatgact accaattgca ttggacttca acaaattgga aaggtaccaa tactaaagat
      901 aaatggacag atcgttcttc agaaagatat aaaatcgatt gggaaaaaga agaaatgaca
      961 aatggcggca gcagtggtgg cagtagtggt ggtgcacaca ttgtgatggt tgatgcgtac
     1021 aaaccgacca agaagggtca tcatcaccat caccattgag tttaaactat atagaataaa
     1081 agaagaaacc ttagctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
     1141 gaggggtttt ttgctgaaag gaggaactat atccggataa ttcttgaaga cgaaagggcc
     1201 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     1261 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     1321 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     1381 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     1441 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     1501 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     1561 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     1621 tattatcccg tgttgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     1681 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     1741 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     1801 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     1861 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     1921 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     1981 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     2041 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     2101 gtgggtcacg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     2161 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     2221 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     2281 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     2341 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     2401 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     2461 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     2521 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     2581 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     2641 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     2701 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     2761 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     2821 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     2881 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     2941 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     3001 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     3061 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     3121 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     3181 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     3241 gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact
     3301 ccgctatcgc tacgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
     3361 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
     3421 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     3481 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     3541 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggccag
     3601 gacccaacgc tgcccgagat ctcgatcccg cgaaat
//
