LOCUS       sequence-196776-        5647 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  K10 peptide with ddFLN4 fingerprint domain and N-terminal ybbr tag 
            for covalent immobilization - resulting in high forces.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5647)
  AUTHORS   Milles LF, Schulten K, Gaub HE, Bernardi RC
  TITLE     Molecular mechanism of extreme mechanostability in a pathogen 
            adhesin.
  JOURNAL   Science. 2018 Mar 30;359(6383):1527-1533. doi: 
            10.1126/science.aar2094.
  PUBMED    29599244
REFERENCE   2  (bases 1 to 5647)
  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: "10.1126/science"; 
            journalName: "Science"; date: "2018-03-30- 30"; volume: "359"; 
            issue: "6383"; pages: "1527-1533"
COMMENT     Sequence Label: pET28a-ybbr-HIS-ddFLN4-K10
FEATURES             Location/Qualifiers
     source          1..5647
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     155..174
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     258..277
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        258..276
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    277..301
                     /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             316..338
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             382..399
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(839..857)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      853..900
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      937..1392
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1024..1043)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1234..1255
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1484..2299)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1484..2299)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2207..2226
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2291..2310
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2421..3009
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2910..2929
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3163..3180
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3195..3334
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3260..3279)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3420..3442
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3436..3627)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4436..5518)
                     /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
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVVVVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5480..5499
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5519..5596)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg
       61 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
      121 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
      181 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga
      241 tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc
      301 ccctgtagaa ataattttgt ttaactttaa gaaggagata tacatatgga ctctctggaa
      361 ttcatcgctt ctaaactggc tcaccaccac caccaccacg gttctgctga cccggaaaaa
      421 tcttacgctg aaggtccggg tctggacggt ggtgaaagtt tccagccgtc taaattcaaa
      481 atccacgctg ttgacccgga cggtgttcac cgtaccgacg gtggtgacgg tttcgttgtt
      541 accatcgaag gtccggctcc ggttgacccg gttatggttg acaacggtga cggtacctac
      601 gacgttgaat tcgaaccgaa agaagctggt gactacgtta tcaacctgac cctggacggt
      661 gacaacgtta acggtttccc gaaaaccgtt accgttaaac cggctccggg ctccggttct
      721 ggttccggct cttacggtgg tggttcttct ggtggtggtt cttctggtgg tggtcactaa
      781 ctcgagtaag atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc
      841 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
      901 ctgaaaggag gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa
      961 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     1021 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     1081 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     1141 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     1201 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     1261 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     1321 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     1381 cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     1441 tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa aactcatcga
     1501 gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa
     1561 gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct
     1621 ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt
     1681 caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg
     1741 gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat
     1801 caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa
     1861 atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga
     1921 acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga
     1981 atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa
     2041 aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat
     2101 ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg
     2161 gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt
     2221 tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag caagacgttt
     2281 cccgttgaat atggctcata acaccccttg tattactgtt tatgtaagca gacagtttta
     2341 ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     2401 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     2461 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     2521 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     2581 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     2641 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     2701 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     2761 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     2821 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     2881 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     2941 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     3001 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     3061 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     3121 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     3181 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     3241 cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     3301 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccaccaa cacccgctga
     3361 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     3421 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
     3481 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
     3541 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
     3601 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
     3661 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
     3721 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
     3781 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
     3841 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
     3901 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
     3961 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
     4021 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
     4081 catgcgcacc cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt
     4141 ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag
     4201 cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag
     4261 cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac
     4321 gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat
     4381 cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact
     4441 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4501 ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg
     4561 gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc
     4621 tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg
     4681 agctgtcttc ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg
     4741 actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag
     4801 tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc
     4861 agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc
     4921 cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct
     4981 ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa
     5041 taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc
     5101 aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac
     5161 tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt
     5221 ctaccaccac caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga
     5281 caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact
     5341 gtttgcccgc cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg
     5401 cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa
     5461 cggtctgata agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat
     5521 tcaccaccct gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc
     5581 gccattcgat ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag
     5641 cagccca
//
