LOCUS       sequence-417178-        5558 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  HaloTag-(Ig32)2-Talin R3-IVVI-(Ig32)2-10xHisTag-AviTag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5558)
  AUTHORS   Tapia-Rojo R, Mora M, Garcia-Manyes S
  TITLE     Single-molecule magnetic tweezers to probe the equilibrium dynamics 
            of individual proteins at physiologically relevant forces and 
            timescales.
  JOURNAL   Nat Protoc. 2024 Mar 11. doi: 10.1038/s41596-024-00965-5.
  PUBMED    38467905
REFERENCE   2  (bases 1 to 5558)
  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 Protoc.
            2024 Mar 11. doi: 10.1038/s41596-024-00965-5."
COMMENT     Sequence Label: pFN18A-HaloTag-Biotin
FEATURES             Location/Qualifiers
     source          1..5558
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..40
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             70..960
                     /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             973..993
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             2488..2517
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     regulatory      2514..2523
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2524..2568
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(2662..2680)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2676..2723
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        2952..3056
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3057..3125,3126..3917)
                     /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(3275..3294)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(4075..4663)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4155..4174)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5122..5139)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5292..5378
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5470..5497
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ggatctcgat cccgcgaaat taatacgact cactataggg tattttaact ttactaagga
       61 gaattcatca tggcagaaat cggtactggc tttccattcg acccccatta tgtggaagtc
      121 ctgggcgagc gcatgcacta cgtcgatgtt ggtccgcgcg atggcacccc tgtgctgttc
      181 ctgcacggta acccgacctc ctcctacgtg tggcgcaaca tcatcccgca tgttgcaccg
      241 acccatcgct gcattgctcc agacctgatc ggtatgggca aatccgacaa accagacctg
      301 ggttatttct tcgacgacca cgtccgcttc atggatgcct tcatcgaagc cctgggtctg
      361 gaagaggtcg tcctggtcat tcacgactgg ggctccgctc tgggtttcca ctgggccaag
      421 cgcaatccag agcgcgtcaa aggtattgca tttatggagt tcatccgccc tatcccgacc
      481 tgggacgaat ggccagaatt tgcccgcgag accttccagg ccttccgcac caccgacgtc
      541 ggccgcaagc tgatcatcga tcagaacgtt tttatcgagg gtacgctgcc gatgggtgtc
      601 gtccgcccgc tgactgaagt cgagatggac cattaccgcg agccgttcct gaatcctgtt
      661 gaccgcgagc cactgtggcg cttcccaaac gagctgccaa tcgccggtga gccagcgaac
      721 atcgtcgcgc tggtcgaaga atacatggac tggctgcacc agtcccctgt cccgaagctg
      781 ctgttctggg gcaccccagg cgttctgatc ccaccggccg aagccgctcg cctggccaaa
      841 agcctgccta actgcaaggc tgtggacatc ggcccgggtc tgaatctgct gcaagaagac
      901 aacccggacc tgatcggcag cgagatcgcg cgctggctgt cgacgctcga gatttccggc
      961 gagccaacca ctgaggatct gtactttcag agcgataacg cgatcgccgg atccgtaatt
     1021 ggtcttctga ggcctttaaa ggatgtcacc gtgactgcag gggaaacagc caccttcgac
     1081 tgcgagctct cctacgaaga tatcccagtg gaatggtatc tcaaagggaa gaaactagag
     1141 cccagcgata aggtggtccc acgttcagaa ggaaaagttc atacacttac tctgagggat
     1201 gtaaagttag aagatgctgg ggaagtccaa ctaacagcaa aagatttcaa aactcacgcc
     1261 aacctctttg tgaaagaacc cgggtccgta attggtcttc tgaggccttt aaaggatgtc
     1321 accgtgactg caggggaaac agccaccttc gactgcgagc tctcctacga agatatccca
     1381 gtggaatggt atctcaaagg gaagaaacta gagcccagcg ataaggtggt cccacgttca
     1441 gaaggaaaag ttcatacact tactctgagg gatgtaaagt tagaagatgc tggggaagtc
     1501 caactaacag caaaagattt caaaactcac gccaacctct ttgtgaaaga accctccgga
     1561 gcccatgcga cgggtgcggg tccagcgggt cgttatgatc aagccaccga caccatcctc
     1621 accgtgatcg agaacatctt cagcagcatg ggtgatgcgg gcgaaatggt tcgccaagcc
     1681 cgcattctgg cccaagcggt tagcgatctg gtgaatgcga tcaaagccga tgcggaaggt
     1741 gaaagcgatc tggagaacag ccgcaaactg ctcagcgccg ccaaaattct ggcggatgcg
     1801 gttgccaaaa tggttgaagc ggccaaaggt gcggccgcgc atccagatag cgaagaacag
     1861 cagcaacgtc tgcgtgaagc ggccgaaggt ctgcgtatgg cgatcaacgc cgcggcgcag
     1921 aatgccatca aaaaggctag cgtaattggt cttctgaggc ctttaaagga tgtcaccgtg
     1981 actgcagggg aaacagccac cttcgactgc gagctctcct acgaagatat cccagtggaa
     2041 tggtatctca aagggaagaa actagagccc agcgataagg tggtcccacg ttcagaagga
     2101 aaagttcata cacttactct gagggatgta aagttagaag atgctgggga agtccaacta
     2161 acagcaaaag atttcaaaac tcacgccaac ctctttgtga aagaacccgg gtccgtaatt
     2221 ggtcttctga ggcctttaaa ggatgtcacc gtgactgcag gggaaacagc caccttcgac
     2281 tgcgagctct cctacgaaga tatcccagtg gaatggtatc tcaaagggaa gaaactagag
     2341 cccagcgata aggtggtccc acgttcagaa ggaaaagttc atacacttac tctgagggat
     2401 gtaaagttag aagatgctgg ggaagtccaa ctaacagcaa aagatttcaa aactcacgcc
     2461 aacctctttg tgaaagaacc cggaggtcac catcaccatc accatcacca tcaccatggt
     2521 ggcggtctga acgacatctt cgaggctcag aaaatcgaat ggcacgaata aggtaccaga
     2581 tcctctagag tcgacctgca ggcatgcaag ctgatccggc tgctaacaaa gcccgaaagg
     2641 aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta
     2701 aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggttcg cttgctgtcc
     2761 ataaaaccgc ccagtctagc tatcgccatg taagcccact gcaagctacc tgctttctct
     2821 ttgcgcttgc gttttccctt gtccagatag cccagtagct gacattcatc cggggtcagc
     2881 accgtttctg cggactggct ttctacgtaa tggtttctta gacgtcaggt ggcacttttc
     2941 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     3001 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     3061 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     3121 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     3181 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     3241 aacgttttcc aatgatgagc actttcaaag ttctgctatg tggcgcggta ttatcccgta
     3301 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     3361 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     3421 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca actatcggag
     3481 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     3541 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     3601 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     3661 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     3721 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     3781 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     3841 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     3901 tgattaagca ttggtaattc gaaatgaccg accaagcgac gcccaaccgg tatcagctca
     3961 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     4021 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     4081 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     4141 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     4201 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     4261 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     4321 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     4381 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     4441 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     4501 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     4561 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     4621 tgtttgcaag cagcagatta cgcgcagaaa aaaaggattt caagaagatc ctttgatctt
     4681 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     4741 attatcaaaa aggatcttca cctagatcct tttatagtct ggaaatacag gaacgcacgc
     4801 tggatggccc ttcgctggga tggtgaaacc atgaaaaatg gcagcttcag tggattaagt
     4861 gggggtaatg tggcctgtac cctctggttg cataggtatt catacggtta aaatttatca
     4921 ggcgcgattg cggcagtttt tcgggtggtt tgttgccatt tttacctgtc tgctgccgtg
     4981 atcgcgctga acgcgtttta gcggtgcgta caattaaggg attatggtaa atccacttac
     5041 tgtctgccct cgtagccatc gagataaacc gcagtactcc ggccacgatg cgtccggcgt
     5101 agaggatcga gatcttttca gcctgataca gattaaatca gaacgcagaa gcggtctgat
     5161 aaaacagaat ttgcctggcg gcagtagcgc ggtggtccca cctgacccca tgccgaactc
     5221 agaagtgaaa cgccgtagcg ccgatggtag tgtggggtct ccccatgcga gagtagggaa
     5281 ctgccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct
     5341 gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc gccgggagcg gatttgaacg
     5401 ttgcgaagca acggcccgga gggtggcggg caggacgccc gccataaact gccaggcatc
     5461 aaattaagca gaaggccatc ctgacggatg gcctttttgc gtttctacaa actcttttgt
     5521 ttatttttct aaatacattc aaatatgtat ccgctcat
//
