LOCUS       sequence-148884-        4965 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Bacterial expression - MBP-his tagged LbCpf1 (E.coli codon 
            optimized).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 4965)
  AUTHORS   Kim D, Kim J, Hur JK, Been KW, Yoon SH, Kim JS
  TITLE     Genome-wide analysis reveals specificities of Cpf1 endonucleases in 
            human cells.
  JOURNAL   Nat Biotechnol. 2016 Jun 6. doi: 10.1038/nbt.3609.
  PUBMED    27272384
REFERENCE   2  (bases 1 to 4965)
  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 
            Biotechnol. 2016 Jun 6. doi: 10.1038/nbt.3609."
COMMENT     Sequence Label: pMAL-his-LbCpf1-EC
FEATURES             Location/Qualifiers
     source          1..4965
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             1..1101
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELVKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1075..1098
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1150..1161
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             1174..1191
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1192..1212
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1231..1251
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1252..4935
                     /codon_start=1
                     /product="CRISPR-associated protein from Lachnospiraceae 
                     bacterium ND2006 (Zetsche et al., 2015)"
                     /label=LbCpf1
                     /translation="MSKLEKFTNCYSLSKTLRFKAIPVGKTQENIDNKRLLVEDEKRAE
                     DYKGVKKLLDRYYLSFINDVLHSIKLKNLNNYISLFRKKTRTEKENKELENLEINLRKE
                     IAKAFKGNEGYKSLFKKDIIETILPEFLDDKDEIALVNSFNGFTTAFTGFFDNRENMFS
                     EEAKSTSIAFRCINENLTRYISNMDIFEKVDAIFDKHEVQEIKEKILNSDYDVEDFFEG
                     EFFNFVLTQEGIDVYNAIIGGFVTESGEKIKGLNEYINLYNQKTKQKLPKFKPLYKQVL
                     SDRESLSFYGEGYTSDEEVLEVFRNTLNKNSEIFSSIKKLEKLFKNFDEYSSAGIFVKN
                     GPAISTISKDIFGEWNVIRDKWNAEYDDIHLKKKAVVTEKYEDDRRKSFKKIGSFSLEQ
                     LQEYADADLSVVEKLKEIIIQKVDEIYKVYGSSEKLFDADFVLEKSLKKNDAVVAIMKD
                     LLDSVKSFENYIKAFFGEGKETNRDESFYGDFVLAYDILLKVDHIYDAIRNYVTQKPYS
                     KDKFKLYFQNPQFMGGWDKDKETDYRATILRYGSKYYLAIMDKKYAKCLQKIDKDDVNG
                     NYEKINYKLLPGPNKMLPKVFFSKKWMAYYNPSEDIQKIYKNGTFKKGDMFNLNDCHKL
                     IDFFKDSISRYPKWSNAYDFNFSETEKYKDIAGFYREVEEQGYKVSFESASKKEVDKLV
                     EEGKLYMFQIYNKDFSDKSHGTPNLHTMYFKLLFDENNHGQIRLSGGAELFMRRASLKK
                     EELVVHPANSPIANKNPDNPKKTTTLSYDVYKDKRFSEDQYELHIPIAINKCPKNIFKI
                     NTEVRVLLKHDDNPYVIGIDRGERNLLYIVVVDGKGNIVEQYSLNEIINNFNGIRIKTD
                     YHSLLDKKEKERFEARQNWTSIENIKELKAGYISQVVHKICELVEKYDAVIALEDLNSG
                     FKNSRVKVEKQVYQKFEKMLIDKLNYMVDKKSNPCATGGALKGYQITNKFESFKSMSTQ
                     NGFIFYIPAWLTSKIDPSTGFVNLLKTKYTSIADSKKFISSFDRIMYVPEEDLFEFALD
                     YKNFSRTDADYIKKWKLYSYGNRIRIFRNPKKNNVFDWEEVCLTSAYKELFNKYGINYQ
                     QGDIRALLCEQSDKAFYSSFMALMSLMLQMRNSITGRTDVDFLISPVKNSDGIFYDSRN
                     YEAQENAILPKNADANGAYNIARKVLWAIGQFKKAEDEKLDKVKIAISNKEWLEYAQTS
                     VKH"
ORIGIN
        1 atgaaaatcg aagaaggtaa actggtaatc tggattaacg gcgataaagg ctataacggt
       61 ctcgctgaag tcggtaagaa attcgagaaa gataccggaa ttaaagtcac cgttgagcat
      121 ccggataaac tggaagagaa attcccacag gttgcggcaa ctggcgatgg ccctgacatt
      181 atcttctggg cacacgaccg ctttggtggc tacgctcaat ctggcctgtt ggctgaaatc
      241 accccggaca aagcgttcca ggacaagctg tatccgttta cctgggatgc cgtacgttac
      301 aacggcaagc tgattgctta cccgatcgct gttgaagcgt tatcgctgat ttataacaaa
      361 gatctgctgc cgaacccgcc aaaaacctgg gaagagatcc cggcgctgga taaagaactg
      421 aaagcgaaag gtaagagcgc gctgatgttc aacctgcaag aaccgtactt cacctggccg
      481 ctgattgctg ctgacggggg ttatgcgttc aagtatgaaa acggcaagta cgacattaaa
      541 gacgtgggcg tggataacgc tggcgcgaaa gcgggtctga ccttcctggt tgacctgatt
      601 aaaaacaaac acatgaatgc agacaccgat tactccatcg cagaagctgc ctttaataaa
      661 ggcgaaacag cgatgaccat caacggcccg tgggcatggt ccaacatcga caccagcaaa
      721 gtgaattatg gtgtaacggt actgccgacc ttcaagggtc aaccatccaa accgttcgtt
      781 ggcgtgctga gcgcaggtat taacgccgcc agtccgaaca aagagctggc aaaagagttc
      841 ctcgaaaact atctgctgac tgatgaaggt ctggaagcgg ttaataaaga caaaccgctg
      901 ggtgccgtag cgctgaagtc ttacgaggaa gagttggtga aagatccgcg tattgccgcc
      961 actatggaaa acgcccagaa aggtgaaatc atgccgaaca tcccgcagat gtccgctttc
     1021 tggtatgccg tgcgtactgc ggtgatcaac gccgccagcg gtcgtcagac tgtcgatgaa
     1081 gccctgaaag acgcgcagac taattcgagc tcgaacaaca acaacaataa caataacaac
     1141 aacctcggga tcgagggaag gatttcacat atgcatcatc atcatcatca tgagaacctg
     1201 tacttccagg gcgaattcgg ctccggacct ccaaagaaaa agagaaaagt aatgagcaaa
     1261 ctggaaaaat ttacgaattg ttatagcctg tccaagaccc tgcgtttcaa agccatcccc
     1321 gttggcaaaa cccaggagaa tattgataat aaacgtctgc tggttgagga tgaaaaaaga
     1381 gcagaagact ataagggagt caaaaaactg ctggatcggt actacctgag ctttataaat
     1441 gacgtgctgc atagcattaa actgaaaaat ctgaataact atattagtct gttccgcaag
     1501 aaaacccgaa cagagaaaga aaataaagag ctggaaaacc tggagatcaa tctgcgtaaa
     1561 gagatcgcaa aagcttttaa aggaaatgaa ggttataaaa gcctgttcaa aaaagacatt
     1621 attgaaacca tcctgccgga atttctggat gataaagacg agatagcgct cgtgaacagc
     1681 ttcaacgggt tcacgaccgc cttcacgggc tttttcgata acagggaaaa tatgttttca
     1741 gaggaagcca aaagcacctc gatagcgttc cgttgcatta atgaaaattt gacaagatat
     1801 atcagcaaca tggatatttt cgagaaagtt gatgcgatct ttgacaaaca tgaagtgcag
     1861 gagattaagg aaaaaattct gaacagcgat tatgatgttg aggatttttt cgagggggaa
     1921 ttttttaact ttgtactgac acaggaaggt atagatgtgt ataatgctat tatcggcggg
     1981 ttcgttaccg aatccggcga gaaaattaag ggtctgaatg agtacatcaa tctgtataac
     2041 caaaagacca aacagaaact gccaaaattc aaaccgctgt acaagcaagt cctgagcgat
     2101 cgggaaagct tgagctttta cggtgaaggt tataccagcg acgaggaggt actggaggtc
     2161 tttcgcaata ccctgaacaa gaacagcgaa attttcagct ccattaaaaa gctggagaaa
     2221 ctgtttaaga attttgacga gtacagcagc gcaggtattt ttgtgaagaa cggacctgcc
     2281 ataagcacca ttagcaagga tatttttgga gagtggaatg ttatccgtga taaatggaac
     2341 gcggaatatg atgacataca cctgaaaaag aaggctgtgg taactgagaa atatgaagac
     2401 gatcgccgca aaagctttaa aaaaatcggc agctttagcc tggagcagct gcaggaatat
     2461 gcggacgccg acctgagcgt ggtcgagaaa ctgaaggaaa ttattatcca aaaagtggat
     2521 gagatttaca aggtatatgg tagcagcgaa aaactgtttg atgcggactt cgttctggaa
     2581 aaaagcctga aaaaaaatga tgctgttgtt gcgatcatga aagacctgct cgatagcgtt
     2641 aagagctttg aaaattacat taaagcattc tttggcgagg gcaaagaaac aaacagagac
     2701 gaaagctttt atggcgactt cgtcctggct tatgacatcc tgttgaaggt agatcatata
     2761 tatgatgcaa ttcgtaatta cgtaacccaa aagccgtaca gcaaagataa gttcaaactg
     2821 tatttccaga acccgcagtt tatgggtggc tgggacaaag acaaggagac agactatcgc
     2881 gccactattc tgcgttacgg cagcaagtac tatctcgcca tcatggacaa aaaatatgca
     2941 aagtgtctgc agaaaatcga taaagacgac gtgaacggaa attacgaaaa gattaattat
     3001 aagctgctgc cagggcccaa caagatgtta ccgaaagtat ttttttccaa aaaatggatg
     3061 gcatactata acccgagcga ggatatacag aagatttaca aaaatgggac cttcaaaaag
     3121 ggggatatgt tcaatctgaa tgactgccac aaactgatcg atttttttaa agatagcatc
     3181 agccgttatc ctaaatggtc aaacgcgtat gattttaatt tctccgaaac ggagaaatat
     3241 aaagacattg ctggtttcta tcgcgaagtc gaagaacagg gttataaagt tagctttgaa
     3301 tcggccagca agaaagaggt tgataaactg gtggaggagg gtaagctgta tatgtttcag
     3361 atttataaca aagactttag cgacaaaagc cacggtactc ctaatctgca tacgatgtac
     3421 tttaaactgc tgtttgatga gaataaccac ggccaaatcc gtctctccgg tggagcagaa
     3481 ctttttatgc ggcgtgcgag cctaaaaaag gaagaactgg tggtgcatcc cgccaacagc
     3541 ccgattgcta acaaaaatcc agataatcct aagaagacca ccacactgtc gtacgatgtc
     3601 tataaggata aacgtttctc ggaagaccag tatgaattgc atataccgat agcaattaat
     3661 aaatgcccaa aaaacatttt caaaatcaac actgaagttc gtgtgctgct gaaacatgat
     3721 gataatccgt atgtgatcgg aattgaccgt ggggagagaa atctgctgta tattgtagtc
     3781 gttgatggca agggcaacat cgttgagcag tatagcctga atgaaataat taataatttt
     3841 aacggtatac gtattaaaac cgactatcat agcctgctgg ataaaaagga gaaagagcgt
     3901 tttgaggcac gccaaaattg gacgagcatc gaaaacatca aggaactgaa ggcaggatat
     3961 atcagccaag tagtccataa aatctgtgaa ctggtggaga agtacgacgc tgtcattgcc
     4021 ctggaagacc tcaatagcgg ctttaaaaac agccgggtga aggtggagaa acaggtatac
     4081 caaaagtttg aaaagatgct cattgataag ctgaactata tggttgataa aaagagcaac
     4141 ccgtgcgcca ctggcggtgc actgaaaggg taccaaatta ccaataaatt tgaaagcttt
     4201 aaaagcatga gcacgcagaa tgggtttatt ttttatatac cagcatggct gacgagcaag
     4261 attgacccca gcactggttt tgtcaatctg ctgaaaacca aatacacaag cattgcggat
     4321 agcaaaaaat ttatttcgag cttcgaccgt attatgtatg ttccggagga agatctgttt
     4381 gaatttgccc tggattataa aaacttcagc cgcaccgatg cagattatat caaaaaatgg
     4441 aagctgtaca gttatggtaa tcgtatacgt atcttccgta atccgaagaa aaacaatgtg
     4501 ttcgattggg aagaggtctg tctgaccagc gcgtataaag aactgttcaa caagtacgga
     4561 ataaattatc agcaaggtga cattcgcgca ctgctgtgtg aacagtcaga taaagcattt
     4621 tatagcagct ttatggcgct gatgagcctg atgctccaga tgcgcaacag cataaccggt
     4681 cgcacagatg ttgactttct gatcagccct gtgaagaata gcgacggcat cttctacgat
     4741 tccaggaact atgaagcaca ggaaaacgct attctgccta aaaatgccga tgccaacggc
     4801 gcctataata ttgcacggaa ggttctgtgg gcgattggac agttcaagaa agcggaagat
     4861 gagaagctgg ataaggtaaa aattgctatt agcaataagg aatggctgga gtacgcacag
     4921 acatcggtta aacacgcggc cgcttccctg caggtaatta aataa
//
