LOCUS       sequence-182540-        4061 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4061)
  AUTHORS   Zakeri B, Fierer JO, Celik E, Chittock EC, Schwarz-Linek U, Moy VT, 
            Howarth M
  TITLE     Peptide tag forming a rapid covalent bond to a protein, through 
            engineering a bacterial adhesin.
  JOURNAL   Proc Natl Acad Sci U S A. 2012 Mar 20;109(12):E690-7. Epub 2012 Feb 
            24.
  PUBMED    22366317
REFERENCE   2  (bases 1 to 4061)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2012-03-20"; volume: "109(12)"; pages: 
            "E690-7. Epub 2012 Feb 24"
COMMENT     Sequence Label: pDEST14-SpyCatcher
FEATURES             Location/Qualifiers
     source          1..4061
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     173..192
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        173..191
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             223..245
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             265..282
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             304..324
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             331..669
                     /codon_start=1
                     /product="truncated version of the CnaB2 adhesin domain of 
                     Streptococcus pyogenes FbaB"
                     /label=SpyCatcher
                     /note="forms a covalent isopeptide bond with the SpyTag 
                     peptide (Zakeri et al., 2012)"
                     /translation="VDTLSGLSSEQGQSGDMTIEEDSATHIKFSKRDEDGKELAGATME
                     LRDSSGKTISTWISDGQVKDFYLYPGKYTFVETAAPDGYEVATAITFTVNEQGQVTVNG
                     KATKGDAHI"
     CDS             734..751
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(804..822)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      818..865
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      902..1357
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(989..1008)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1199..1220
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1383..1487
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1488..1556,1557..2348)
                     /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(1706..1725)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2519..3107
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3008..3027
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3261..3278
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3293..3432
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3358..3377)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3518..3540
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3534..3725)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg
       61 ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat
      121 gcgcacccgt ggccaggacc caacgctgcc cgagatctcg atcccgcgaa attaatacga
      181 ctcactatag ggagaccaca acggtttccc tctagaaata attttgttta actttaagaa
      241 ggagatatac atatgtcgta ctaccatcac catcaccatc acgattacga catcccaacg
      301 accgaaaacc tgtattttca gggcgccatg gttgatacct tatcaggttt atcaagtgag
      361 caaggtcagt ccggtgatat gacaattgaa gaagatagtg ctacccatat taaattctca
      421 aaacgtgatg aggacggcaa agagttagct ggtgcaacta tggagttgcg tgattcatct
      481 ggtaaaacta ttagtacatg gatttcagat ggacaagtga aagatttcta cctgtatcca
      541 ggaaaatata catttgtcga aaccgcagca ccagacggtt atgaggtagc aactgctatt
      601 acctttacag ttaatgagca aggtcaggtt actgtaaatg gcaaagcaac taaaggtgac
      661 gctcatattt aaatggttga tgcttgagga tccgaattcg agctccgtcg acaagcttgc
      721 ggccgcactc gagcaccacc accaccacca ctgagatccg gctgctaaca aagcccgaaa
      781 ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc
      841 taaacgggtc ttgaggggtt ttttgctgaa aggaggaact atatccggat tggcgaatgg
      901 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
      961 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     1021 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     1081 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     1141 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     1201 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     1261 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     1321 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     1381 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     1441 gacaataacc ctggtaaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     1501 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     1561 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     1621 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     1681 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     1741 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     1801 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     1861 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     1921 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     1981 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg
     2041 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     2101 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     2161 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     2221 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     2281 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     2341 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     2401 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     2461 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     2521 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     2581 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     2641 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     2701 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     2761 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     2821 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     2881 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     2941 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     3001 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     3061 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     3121 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     3181 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     3241 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc
     3301 ggtattttct ccttacgcat ctgtgcggta tttcacaccg caatggtgca ctctcagtac
     3361 aatctgctct gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg
     3421 gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     3481 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     3541 ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg
     3601 tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca gctcgttgag cttctccaga
     3661 agcgttaatg tctggcttct gataaagcgg gccatgttaa gggcggtttt ttcctgtttg
     3721 gtcactgatg cctccgtgta agggggattt ctgttcatgg gggtaatgat accgatgaaa
     3781 cgagagagga tgctcacgat acgggttact gatgatgaac atgcccggtt actggaacgt
     3841 tgtgagggta aacaactggc ggtatggatg cggcgggacc agagaaaaat cactcagggt
     3901 caatgccagc gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct
     3961 gcgatgcaga tccggaacat aatggtgcag ggcgctgact tccgcgtttc cagactttac
     4021 gaaacacgga aaccgaagac cattcatgtt gttgctcagg t
//
