LOCUS       Exported                5829 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Encodes Cas9 as a Type 3 part to be used in the Dueber YTK system.
ACCESSION   .
VERSION     .
KEYWORDS    pYTK036
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5829)
  AUTHORS   Lee ME, DeLoache WC, Cervantes B, Dueber JE
  TITLE     A Highly-characterized Yeast Toolkit for Modular, Multi-part 
            Assembly.
  JOURNAL   ACS Synth Biol. 2015 Apr 14.
  PUBMED    25871405
REFERENCE   2  (bases 1 to 5829)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5829
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             11..4114
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             4127..4147
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4151..4171
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      4194..4228
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     CDS             complement(4299..4958)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4873..4892
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     rep_origin      5140..5728
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5629..5648
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcggtctcct atggacaaga agtattctat cggactggat atcgggacta atagcgtcgg
       61 gtgggccgtg atcactgacg agtacaaggt gccctctaag aagttcaagg tgctcgggaa
      121 caccgaccgg cattccatca agaaaaatct gatcggagct ctcctctttg attcagggga
      181 aaccgctgaa gcaacccgcc tcaagcggac tgctagacgg cggtacacca ggaggaagaa
      241 ccggatttgt taccttcaag agatattctc caacgaaatg gcaaaggtcg acgacagctt
      301 cttccatagg ctggaagaat cattcctcgt ggaagaggat aagaagcatg aacggcatcc
      361 catcttcggt aatatcgtcg acgaggtggc ctatcacgag aaatacccaa ccatctacca
      421 tcttcgcaaa aagctggtgg actcaaccga caaggcagac ctccggctta tctacctggc
      481 cctggcccac atgatcaagt tcagaggcca cttcctgatc gagggcgacc tcaatcctga
      541 caatagcgat gtggataaac tgttcatcca gctggtgcag acttacaacc agctctttga
      601 agagaacccc atcaatgcaa gcggagtcga tgccaaggcc attctgtcag cccggctgtc
      661 aaagagccgc agacttgaga atcttatcgc tcagctgccg ggtgaaaaga aaaatggact
      721 gttcgggaac ctgattgctc tttcacttgg gctgactccc aatttcaagt ctaatttcga
      781 cctggcagag gatgccaagc tgcaactgtc caaggacacc tatgatgacg atctcgacaa
      841 cctcctggcc cagatcggtg accaatacgc cgaccttttc cttgctgcta agaatctttc
      901 tgacgccatc ctgctgtctg acattctccg cgtgaacact gaaatcacca aggcccctct
      961 ttcagcttca atgattaagc ggtatgatga gcaccaccag gacctgaccc tgcttaaggc
     1021 actcgtccgg cagcagcttc cggagaagta caaggaaatc ttctttgacc agtcaaagaa
     1081 tggatacgcc ggctacatcg acggaggtgc ctcccaagag gaattttata agtttatcaa
     1141 acctatcctt gagaagatgg acggcaccga agagctcctc gtgaaactga atcgggagga
     1201 tctgctgcgg aagcagcgca ctttcgacaa tgggagcatt ccccaccaga tccatcttgg
     1261 ggagcttcac gccatccttc ggcgccaaga ggacttctac ccctttctta aggacaacag
     1321 ggagaagatt gagaaaattc tcactttccg catcccctac tacgtgggac ccctcgccag
     1381 aggaaatagc cggtttgctt ggatgaccag aaagtcagaa gaaactatca ctccctggaa
     1441 cttcgaagag gtggtggaca agggagccag cgctcagtca ttcatcgaac ggatgactaa
     1501 cttcgataag aacctcccca atgagaaggt cctgccgaaa cattccctgc tctacgagta
     1561 ctttaccgtg tacaacgagc tgaccaaggt gaaatatgtc accgaaggga tgaggaagcc
     1621 cgcattcctg tcaggcgaac aaaagaaggc aattgtggac cttctgttca agaccaatag
     1681 aaaggtgacc gtgaagcagc tgaaggagga ctatttcaag aaaattgaat gcttcgactc
     1741 tgtggagatt agcggggtcg aagatcggtt caacgcaagc ctgggtactt accatgatct
     1801 gcttaagatc atcaaggaca aggattttct ggacaatgag gagaacgagg acatccttga
     1861 ggacattgtc ctgactctca ctctgttcga ggaccgggaa atgatcgagg agaggcttaa
     1921 gacctacgcc catctgttcg acgataaagt gatgaagcaa cttaaacgga gaagatatac
     1981 cggatgggga cgccttagcc gcaaactcat caacggaatc cgggacaaac agagcggaaa
     2041 gaccattctt gatttcctta agagcgacgg attcgctaat cgcaacttca tgcaacttat
     2101 ccatgatgat tccctgacct ttaaggagga catccagaag gcccaagtgt ctggacaagg
     2161 tgactcactg cacgagcata tcgcaaatct ggctggttca cccgctatta agaagggtat
     2221 tctccagacc gtgaaagtcg tggacgagct ggtcaaggtg atgggtcgcc ataaaccaga
     2281 gaacattgtc atcgagatgg ccagggaaaa ccagactacc cagaagggac agaagaacag
     2341 cagggagcgg atgaaaagaa ttgaggaagg gattaaggag ctcgggtcac agatccttaa
     2401 agagcacccg gtggaaaaca cccagcttca gaatgagaag ctctatctgt actaccttca
     2461 aaatggacgc gatatgtatg tggaccaaga gcttgatatc aacaggctct cagactacga
     2521 cgtggaccat atcgtccctc agagcttcct caaagacgac tcaattgaca ataaggtgct
     2581 gactcgctca gacaagaacc ggggaaagtc agataacgtg ccctcagagg aagtcgtgaa
     2641 aaagatgaag aactattggc gccagcttct gaacgcaaag ctgatcactc agcggaagtt
     2701 cgacaatctc actaaggctg agaggggcgg actgagcgaa ctggacaaag caggattcat
     2761 taaacggcaa cttgtggaga ctcggcagat tactaaacat gtcgcccaaa tccttgactc
     2821 acgcatgaat accaagtacg acgaaaacga caaacttatc cgcgaggtga aggtgattac
     2881 cctgaagtcc aagctggtca gcgatttcag aaaggacttt caattctaca aagtgcggga
     2941 gatcaataac tatcatcatg ctcatgacgc atatctgaat gccgtggtgg gaaccgccct
     3001 gatcaagaag tacccaaagc tggaaagcga gttcgtgtac ggagactaca aggtctacga
     3061 cgtgcgcaag atgattgcca aatctgagca ggagatcgga aaggccaccg caaagtactt
     3121 cttctacagc aacatcatga atttcttcaa gaccgaaatc acccttgcaa acggtgagat
     3181 ccggaagagg ccgctcatcg agactaatgg ggagactggc gaaatcgtgt gggacaaggg
     3241 cagagatttc gctaccgtgc gcaaagtgct ttctatgcct caagtgaaca tcgtgaagaa
     3301 aaccgaggtg caaaccggag gcttttctaa ggaatcaatc ctccccaagc gcaactccga
     3361 caagctcatt gcaaggaaga aggattggga ccctaagaag tacggcggat tcgattcacc
     3421 aactgtggct tattctgtcc tggtcgtggc taaggtggaa aaaggaaagt ctaagaagct
     3481 caagagcgtg aaggaactgc tgggtatcac cattatggag cgcagctcct tcgagaagaa
     3541 cccaattgac tttctcgaag ccaaaggtta caaggaagtc aagaaggacc ttatcatcaa
     3601 gctcccaaag tatagcctgt tcgaactgga gaatgggcgg aagcggatgc tcgcctccgc
     3661 tggcgaactt cagaagggta atgagctggc tctcccctcc aagtacgtga atttcctcta
     3721 ccttgcaagc cattacgaga agctgaaggg gagccccgag gacaacgagc aaaagcaact
     3781 gtttgtggag cagcataagc attatctgga cgagatcatt gagcagattt ccgagttttc
     3841 taaacgcgtc attctcgctg atgccaacct cgataaagtc cttagcgcat acaataagca
     3901 cagagacaaa ccaattcggg agcaggctga gaatatcatc cacctgttca ccctcaccaa
     3961 tcttggtgcc cctgccgcat tcaagtactt cgacaccacc atcgaccgga aacgctatac
     4021 ctccaccaaa gaagtgctgg acgccaccct catccaccag agcatcaccg gactttacga
     4081 aactcggatt gacctctcac agctcggagg ggatgaggga gctgatccaa aaaagaagag
     4141 aaaggtagat ccaaaaaaga agagaaaggt ataaggatcc tgagaccaga ccaataaaaa
     4201 acgcccggcg gcaaccgagc gttctgaaca aatccagatg gagttctgag gtcattactg
     4261 gatctatcaa caggagtcca agcgagctcg atatcaaatt acgccccgcc ctgccactca
     4321 tcgcagtact gttgtaattc attaagcatt ctgccgacat ggaagccatc acaaacggca
     4381 tgatgaacct gaatcgccag cggcatcagc accttgtcgc cttgcgtata atatttgccc
     4441 atggtgaaaa cgggggcgaa gaagttgtcc atattggcca cgtttaaatc aaaactggtg
     4501 aaactcaccc agggattggc tgaaacgaaa aacatattct caataaaccc tttagggaaa
     4561 taggccaggt tttcaccgta acacgccaca tcttgcgaat atatgtgtag aaactgccgg
     4621 aaatcgtcgt ggtattcact ccagagcgat gaaaacgttt cagtttgctc atggaaaacg
     4681 gtgtaacaag ggtgaacact atcccatatc accagctcac cgtctttcat tgccatacga
     4741 aattccggat gagcattcat caggcgggca agaatgtgaa taaaggccgg ataaaacttg
     4801 tgcttatttt tctttacggt ctttaaaaag gccgtaatat ccagctgaac ggtctggtta
     4861 taggtacatt gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat
     4921 atatcaacgg tggtatatcc agtgattttt ttctccattt tagcttcctt agctcctgaa
     4981 aatctcgata actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg
     5041 gaacctctta cgtgcccgat caatcatgac caaaatccct taacgtgagt tttcgttcca
     5101 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5161 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     5221 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     5281 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     5341 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     5401 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5461 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     5521 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     5581 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     5641 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     5701 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     5761 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     5821 taaccgtag
//
