LOCUS       sequence-375048-        4997 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid expresses human talin R7R8 domain flanked between a HaloTag 
            and a SpyTag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4997)
  AUTHORS   Dahal N, Sharma S, Phan B, Eis A, Popa I
  TITLE     Mechanical regulation of talin through binding and history-dependent
            unfolding.
  JOURNAL   Sci Adv. 2022 Jul 15;8(28):eabl7719. doi: 10.1126/sciadv.abl7719. 
            Epub 2022 Jul 15.
  PUBMED    35857491
REFERENCE   2  (bases 1 to 4997)
  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: "Sci Adv."; 
            date: "2022-07-15"; volume: "8(28):eabl7719. doi"; pages: " 
            10.1126/sciadv.abl7719. Epub 2022 Jul 15"
COMMENT     Sequence Label: pFN-HaloTag-TalinR7R8-SpyTag
FEATURES             Location/Qualifiers
     source          1..4997
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      41..68
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     150..169
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        150..168
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             199..1089
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             1102..1122
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             2062..2100
                     /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             2116..2133
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2230..2248)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2244..2291
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        2520..2624
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2625..2693,2694..3485)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2843..2862)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(3643..4231)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3723..3742)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    4347..4630
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     primer_bind     complement(4690..4707)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      4860..4946
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gcaggacgcc cgccataaac tgccaggcat caaattaagc agaaggccat cctgacggat
       61 ggcctttttg cgtttctaca aactcttttg tttatttttc taaatacatt caaatatgta
      121 tccgctcatg gatctcgatc ccgcgaaatt aatacgactc actatagggt attttaactt
      181 tactaaggag aattcatcat ggcagaaatc ggtactggct ttccattcga cccccattat
      241 gtggaagtcc tgggcgagcg catgcactac gtcgatgttg gtccgcgcga tggcacccct
      301 gtgctgttcc tgcacggtaa cccgacctcc tcctacgtgt ggcgcaacat catcccgcat
      361 gttgcaccga cccatcgctg cattgctcca gacctgatcg gtatgggcaa atccgacaaa
      421 ccagacctgg gttatttctt cgacgaccac gtccgcttca tggatgcctt catcgaagcc
      481 ctgggtctgg aagaggtcgt cctggtcatt cacgactggg gctccgctct gggtttccac
      541 tgggccaagc gcaatccaga gcgcgtcaaa ggtattgcat ttatggagtt catccgccct
      601 atcccgacct gggacgaatg gccagaattt gcccgcgaga ccttccaggc cttccgcacc
      661 accgacgtcg gccgcaagct gatcatcgat cagaacgttt ttatcgaggg tacgctgccg
      721 atgggtgtcg tccgcccgct gactgaagtc gagatggacc attaccgcga gccgttcctg
      781 aatcctgttg accgcgagcc actgtggcgc ttcccaaacg agctgccaat cgccggtgag
      841 ccagcgaaca tcgtcgcgct ggtcgaagaa tacatggact ggctgcacca gtcccctgtc
      901 ccgaagctgc tgttctgggg caccccaggc gttctgatcc caccggccga agccgctcgc
      961 ctggccaaaa gcctgcctaa ctgcaaggct gtggacatcg gcccgggtct gaatctgctg
     1021 caagaagaca acccggacct gatcggcagc gagatcgcgc gctggctgtc gacgctcgag
     1081 atttccggcg agccaaccac tgaggatctg tactttcaga gcgataacgc gatcgccgga
     1141 tccatgcagc aggcaccggg ccaaaaagaa tgtgacaatg ccctgcgcga actggagacc
     1201 gtgcgcgaac tgctggagaa tccggtgcag ccgatcaatg atatgagtta tttcggctgc
     1261 ctggacagtg tgatggagaa cagcaaagtg ctgggtgagg ccatgaccgg tatcagccag
     1321 aacgccaaaa acggcaatct gccggaattc ggcgatgcca tcagcacagc aagcaaagcc
     1381 ctgtgcggct ttaccgaagc agccgcccag gcagcatatc tggtgggtgt gagcgatccg
     1441 aatagccagg ccggtcaaca gggtctggtt gaaccgaccc agtttgcccg cgccaatcag
     1501 gccatccaga tggcatgcca gagcttaggt gagccgggtt gcacccaggc ccaggttctg
     1561 agcgccgcaa ccattgttgc caaacatacc agcgccctgt gtaatagctg ccgcctggca
     1621 agcgcccgta ccacaaatcc gaccgccaaa cgccagttcg tgcagagcgc caaagaagtg
     1681 gccaatagca cagccaacct ggttaaaacc atcaaggccc tggacggtgc atttaccgag
     1741 gaaaatcgcg cacagtgtcg cgcagcaaca gccccgctgc tggaagcagt tgacaacctg
     1801 agcgccttcg ccagcaatcc tgagtttagc agcattccgg cacagattag cccggagggc
     1861 cgcgcagcaa tggagccgat tgtgatcagc gccaaaacaa tgctggaaag cgccggcggc
     1921 ttaatccaga cagcccgtgc cctggccgtg aatcctcgcg atccgccgag ctggagtgtt
     1981 ctggccggtc atagccgtac cgtgagcgac agcatcaaga agctgatcac aagcatgcgc
     2041 gataaagcac cgggtagatc cgcccacatc gtgatggtgg acgcctacaa gccgacgaag
     2101 gggggttgta gatcccacca tcaccatcac cattgctaag gtaccagatc ctctagagtc
     2161 gacctgcagg catgcaagct gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     2221 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     2281 ggggtttttt gctgaaagga ggaactatat ccggttcgct tgctgtccat aaaaccgccc
     2341 agtctagcta tcgccatgta agcccactgc aagctacctg ctttctcttt gcgcttgcgt
     2401 tttcccttgt ccagatagcc cagtagctga cattcatccg gggtcagcac cgtttctgcg
     2461 gactggcttt ctacgtaatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     2521 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     2581 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     2641 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     2701 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     2761 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     2821 tgatgagcac tttcaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     2881 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     2941 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     3001 ccatgagtga taacactgcg gccaacttac ttctgacaac tatcggagga ccgaaggagc
     3061 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     3121 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     3181 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     3241 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     3301 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     3361 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     3421 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     3481 ggtaattcga aatgaccgac caagcgacgc ccaaccggta tcagctcact caaaggcggt
     3541 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     3601 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     3661 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     3721 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     3781 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     3841 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     3901 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3961 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4021 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     4081 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     4141 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     4201 gcagattacg cgcagaaaaa aaggatttca agaagatcct ttgatctttt ctacggggtc
     4261 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     4321 gatcttcacc tagatccttt tatagtccgg aaatacagga acgcacgctg gatggccctt
     4381 cgctgggatg gtgaaaccat gaaaaatggc agcttcagtg gattaagtgg gggtaatgtg
     4441 gcctgtaccc tctggttgca taggtattca tacggttaaa atttatcagg cgcgattgcg
     4501 gcagtttttc gggtggtttg ttgccatttt tacctgtctg ctgccgtgat cgcgctgaac
     4561 gcgttttagc ggtgcgtaca attaagggat tatggtaaat ccacttactg tctgccctcg
     4621 tagccatcga gataaaccgc agtactccgg ccacgatgcg tccggcgtag aggatcgaga
     4681 tcttttcagc ctgatacaga ttaaatcaga acgcagaagc ggtctgataa aacagaattt
     4741 gcctggcggc agtagcgcgg tggtcccacc tgaccccatg ccgaactcag aagtgaaacg
     4801 ccgtagcgcc gatggtagtg tggggtctcc ccatgcgaga gtagggaact gccaggcatc
     4861 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
     4921 tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt gcgaagcaac
     4981 ggcccggagg gtggcgg
//
