LOCUS       sequence-306487-        6595 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of Rc. CohE wild-type construct containing ybbr 
            tag, ELP linker and ddFLN4 fingerprint domain. This construct was 
            designed for AFM measurements..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6595)
  AUTHORS   Liu Z, Liu H, Vera AM, Bernardi RC, Tinnefeld P, Nash MA
  TITLE     High force catch bond mechanism of bacterial adhesion in the human 
            gut.
  JOURNAL   Nat Commun. 2020 Aug 28;11(1):4321. doi: 10.1038/s41467-020-18063-x.
  PUBMED    32859904
REFERENCE   2  (bases 1 to 6595)
  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: "2020-08-28"; volume: "11(1):4321. doi"; pages: " 
            10.1038/s41467-020-18063-x"
COMMENT     Sequence Label: pET28a-Coh-ddFLN4-ELP-HIS-ybbr
FEATURES             Location/Qualifiers
     source          1..6595
                     /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"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1412..1429
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1530..1548)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1544..1591
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1628..2083
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1715..1734)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1925..1946
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2175..2990)
                     /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(2175..2990)
                     /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     2898..2917
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2982..3001
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3112..3700
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3601..3620
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3854..3871
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3886..4025
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3951..3970)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4111..4133
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4127..4318)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5127..6209)
                     /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     6171..6190
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6210..6287)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6493..6512
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctgta gaaataattt
       61 tgtttaactt taagaaggag atatacatat ggctgacggt gcagctaagc ttagtatgga
      121 ccaaaaattc gctgagcctg gtgaaacggt ggagatcgca ctgaatttgg agaactttga
      181 cgccagctgg acgggtttag agtttttggt caactacgac ccaaaactgg aggtcgcgct
      241 tgatggagct ggagatattg attattccta tggcgacgcc attggcgcta tgggtaagaa
      301 aatcagcgtc ggaggggcga tctccaagga cttgaccgct gatggtctta aagggtttgc
      361 ctttgcctgg ggtactgcca cggcaatctc aggcaacggg caattaggtg tgtttaagtt
      421 taccgttcca gcggatgctc aaccggggga tgagtttcct gttaatttaa ctgtgaacgt
      481 cgggtcgttt attgacgcca acaaggaaaa catcccgttt gaaaccgtaa atggctggat
      541 caaaattaag gaagagggct ccggttctgg atccggttct gctgacccgg aaaaatctta
      601 cgctgaaggt ccgggtctgg acggtggtga atgcttccag ccgtctaaat tcaaaatcca
      661 cgctgttgac ccggacggtg ttcaccgtac cgacggtggt gacggtttcg ttgttaccat
      721 cgaaggtccg gctccggttg acccggttat ggttgacaac ggtgacggta cctacgacgt
      781 tgaattcgaa ccgaaagaag ctggtgacta cgttatcaac ctgaccctgg acggtgacaa
      841 cgttaacggt ttcccgaaaa ccgttaccgt taaaccggct ccgggctccg gttctggttc
      901 ccacggcgtg ggtgttccag gtatgggtgt gcctggtgtt ggtgttcctg gcgttggcgt
      961 gccaggtgtg ggagtaccag gcgtaggcgt tcccggtgta ggcgtgcctg gcgtcggtgt
     1021 acccggtgtc ggagtccctg gtgaaggtgt gccaggcgaa ggtgtgccgg gcgtgggtgt
     1081 tccaggtatg ggtgtgcctg gtgttggtgt tcctggcgtt ggcgtgccag gtgtgggagt
     1141 accaggcgta ggcgttcccg gtgtaggcgt gcctggcgtc ggtgtacccg gtgtcggagt
     1201 ccctggtgaa ggtgtgccag gcgaaggtgt gccgggcgtg ggtgttccag gtatgggtgt
     1261 gcctggtgtt ggtgttcctg gcgttggcgt gccaggtgtg ggagtaccag gcgtaggcgt
     1321 tcccggtgta ggcgtgcctg gcgtcggtgt acccggtgtc ggagtccctg gtgaaggtgt
     1381 gccaggcgaa ggtgtgccgg gctggcgcgg tcatcatcat caccatcatg gtagcgactc
     1441 tctggaattc atcgcttcta aactggctta actcgagtaa gatccggctg ctaacaaagc
     1501 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1561 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     1621 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1681 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1741 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1801 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1861 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1921 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1981 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2041 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     2101 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2161 tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata
     2221 tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca
     2281 ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca
     2341 acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca
     2401 ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact
     2461 tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta
     2521 ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta
     2581 caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca
     2641 cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg
     2701 agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat
     2761 tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg
     2821 ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca
     2881 cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg
     2941 gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt
     3001 gtattactgt ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga
     3061 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     3121 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     3181 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3241 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3301 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3361 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3421 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3481 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     3541 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     3601 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     3661 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     3721 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     3781 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     3841 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     3901 tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc
     3961 tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg
     4021 ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     4081 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     4141 cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg
     4201 attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta
     4261 atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg
     4321 atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga
     4381 ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg
     4441 gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag ggtcaatgcc
     4501 agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat cctgcgatgc
     4561 agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt tacgaaacac
     4621 ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc
     4681 ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa ccccgccagc
     4741 ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc gccatgccgg
     4801 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
     4861 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
     4921 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
     4981 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     5041 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     5101 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     5161 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     5221 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     5281 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     5341 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     5401 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     5461 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     5521 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     5581 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     5641 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     5701 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     5761 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     5821 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     5881 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     5941 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     6001 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     6061 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     6121 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     6181 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     6241 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     6301 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     6361 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     6421 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     6481 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     6541 gatgccggcc acgatgcgtc cggcgtagag gatcgagatc tcgatcccgc gaaat
//
