LOCUS Exported 14200 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Clostridium difficile CRISPR-cas9 mutagenesis plasmid Tn916 oriT. ACCESSION . VERSION . KEYWORDS pKM126 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 14200) AUTHORS McAllister KN, Bouillaut L, Kahn JN, Self WT, Sorg JA TITLE Using CRISPR-Cas9-mediated genome editing to generate C. difficile mutants defective in selenoproteins synthesis. JOURNAL Sci Rep. 2017 Nov 7;7(1):14672. doi: 10.1038/s41598-017-15236-5. PUBMED 29116155 REFERENCE 2 (bases 1 to 14200) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..14200 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 63..79 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 63..79 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" CDS complement(2117..2737) /codon_start=1 /gene="tetR from transposon Tn10" /product="tetracycline repressor TetR" /label=TetR /note="TetR binds to the tetracycline operator tetO to inhibit transcription. This inhibition can be relieved by adding tetracycline or doxycycline." /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS" primer_bind 2645..2664 /label=Tet-R /note="Tetracycline resistance gene, reverse primer" protein_bind 2847..2865 /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O1 for the tetR and tetA genes" CDS 2938..7044 /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" primer_bind complement(7054..7071) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(7054..7070) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(7063..7085) /label=M13/pUC Forward /note="In lacZ gene" misc_RNA 12344..12419 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" rep_origin 12617..13205 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 13106..13125 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 13359..13376 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 cctgcaggat aaaaaaattg tagataaatt ttataaaata gttttatcta caattttttt 61 atcaggaaac agctatgacc gcggccgcga cgtgattttt aatgggtaat ttaagtgtga 121 aatttggaag tgtagaattt aaaaatccag taataatggc gtctggaact tttggatttg 181 gaaaagaata taatgagatt tatgatatac aaaaacttgg tggtataagt agcaagggtc 241 ttactttaaa taaaaaacct ggtaacaatg gtatgagagt tcatgagaca ccatccggta 301 tgatgaatag tgttgggctt gaaaaccctg gagttcaagg atttatagac tatgaattac 361 ctttttttag taaactagac ttagtcagaa tagctaatgt tggaggtggg acattagaag 421 actacttatt aggggttcaa atgttaaatg acaagcctat agacattatt gaattaaata 481 tatcttgccc aaatgtcaaa gctggtggaa tggcttttgg tataaaaaat gaagtggcaa 541 gagaagttgt tcgtgaggta agaaacataa cgaaactgcc tttagtaata aaattatcac 601 ccaatgcaga agatattgta ggaatggcaa aagtttgtga agaagaaggt gctgatggag 661 tttcattagt taatactttt aaagcaatgg caattgatat aaaaaataga agaccagtat 721 ttgaaaatgt atatgctgga ttatctggac cagctataaa acctatagct ttgagaatgg 781 tacatgaagt gtgtaaaaat gtaaatatac ctgtaatggg aatgggcgga ataactaaag 841 ccacagatgc tatagagttt ataatggcag gagctacttg tatccaagtt ggaactgcaa 901 attttatgaa tcctagaata ggtattgaaa taatagatgg gattaatgaa tttatggata 961 gagaaggaat aaaaagttta gacgaaataa gaggaattat ataaggtaga ataaatagta 1021 ttttaaagat aaaaaatata gattaaataa acaataattt tataacatta acatggagga 1081 aaaataatta aaaacttaat tatttatagt gttacttaaa aaatgtaaat tttttagagt 1141 aaaaatttgt aatataacat ataacttaaa aataaaaagc agttagcctt ataaatgaag 1201 gttagctgtt ttttttgtta aaaatatatt tacaaaataa gagatagtag cagtacaata 1261 taagtataag gtaaaagtat tatatattag ggaggggtaa gtatgaaaaa tcaattaaat 1321 gaaaatcata aaaaggttgt attttttgga gtaggtgctg taggagctac ttttgcagaa 1381 caatttttta actctaaata tgattttaaa attctttgtg ataatgaaag aaagaaaaga 1441 tatttagaag aaggatttat aataaatgga aaaaggtatg attttaatta tgtaactaaa 1501 gatgagtata aacaagaggc tgattttata attataggtc taaaatataa taatttaaaa 1561 gaaaatataa aagaattaga tggattagtt ggaaaaaata cagttataat gtctctgcta 1621 aatggagttg atagcgaaga gataatagga gaaagatttg gaattggaaa aatggtatat 1681 tcatatgtta ccaatataga tgcaaagaaa atcaataata atattataca cactactaat 1741 gggataattg tatttggtaa caaagataat agtgaagata gaaaaactaa tataataact 1801 gaagtctttg atgatgtaaa tatagaatat attttatcaa aagatattca acgagatatg 1861 tggtggaagt acatggttaa tattggtgta aatcaaactt cagctatact tggtgcacct 1921 tatggagttt ttcagagttc tgagcattta agagaattag caaaatctgc aatgagggaa 1981 gttgttgcta tagcacaagc aaaagacata tctcttacag aagatgatgt tgaaaattca 2041 ttacatagaa tactagaaca ttcaaaagaa ggaagaacat caatgcttct cgaggcctgc 2101 agacatgcgc gatcgcagac ccactttcac atttaagttg tttttctaat ccgcatatga 2161 tcaattcaag gccgaataag aaggctggct ctgcaccttg gtgatcaaat aattcgatag 2221 cttgtcgtaa taatggcggc atactatcag tagtaggtgt ttccctttct tctttagcga 2281 cttgatgctc ttgatcttcc aatacgcaac ctaaagtaaa atgccccaca gcgctgagtg 2341 catataatgc attctctagt gaaaaacctt gttggcataa aaaggctaat tgattttcga 2401 gagtttcata ctgtttttct gtaggccgtg tacctaaatg tacttttgct ccatcgcgat 2461 gacttagtaa agcacatcta aaacttttag cgttattacg taaaaaatct tgccagcttt 2521 ccccttctaa agggcaaaag tgagtatggt gcctatctaa catctcaatg gctaaggcgt 2581 cgagcaaagc ccgcttattt tttacatgcc aatacaatgt aggctgctct acacctagct 2641 tctgggcgag tttacgggtt gttaaacctt cgattccgac ctcattaagc agctctaatg 2701 cgctgttaat cactttactt ttatctaatc tagacatcat taattcctcc tttttgttga 2761 cattatatca ttgatagagt tatttgtcaa actagttttt tatttcgatg ccctggactt 2821 catgaaaaac taaaaaaaat attgacactc tatcattgat agagtataat taaaataagc 2881 ttgatcgtag cgttaacaga tctgagctca ataatactag gaggtttttt taattaaatg 2941 gataaaaaat atagtatagg attagatata ggaacaaata gtgtaggatg ggcagtaata 3001 actgatgaat ataaagtacc aagtaaaaaa tttaaagtat taggaaatac agatagacat 3061 agtataaaaa aaaatttaat aggtgcatta ctttttgata gtggtgaaac agcagaagca 3121 acaagactta aaagaacagc aagaagaaga tatactagaa gaaaaaatag aatatgttat 3181 ttacaagaaa tatttagtaa tgaaatggca aaagtagatg atagtttttt tcatagatta 3241 gaagaaagtt ttttagtaga agaagataaa aaacacgaaa gacatccaat atttggaaat 3301 atagtagatg aagtagcata tcatgaaaaa tatccaacaa tatatcattt aagaaaaaaa 3361 cttgtagata gtacagataa agcagattta agacttatat atcttgcatt agcacatatg 3421 ataaaattta gaggacattt tcttatagaa ggtgatttaa atccagataa tagtgatgta 3481 gataaacttt ttatacaatt agtacaaaca tataatcaat tatttgaaga aaatccaata 3541 aatgcaagtg gtgtagatgc aaaagcaata ttaagtgcaa gattaagtaa aagtagaaga 3601 ttagaaaatc ttatagcaca attacctgga gaaaaaaaaa atggattatt tggtaatctt 3661 atagctttaa gtttaggatt aacaccaaat tttaaatcaa attttgatct tgcagaagat 3721 gcaaaacttc aattaagtaa agatacatat gatgatgatt tagataattt acttgcacaa 3781 ataggtgatc aatatgcaga tttatttctt gcagcaaaaa atctttctga tgcaatactt 3841 ttaagtgata tacttagagt aaatacagaa ataacaaaag caccattatc tgcaagtatg 3901 ataaaaagat atgatgaaca tcatcaagat ttaacattac ttaaagcatt agtaagacaa 3961 caattaccag aaaaatataa agaaattttt tttgatcaaa gtaaaaatgg atatgcagga 4021 tatatagatg gtggtgcaag tcaagaagaa ttttataaat ttattaaacc aatattagaa 4081 aaaatggatg gaacagaaga attattagta aaacttaata gagaagattt acttagaaaa 4141 caaagaacat ttgataatgg aagtatacca catcaaatac atttaggtga attacatgca 4201 atattaagaa gacaagaaga tttttatcca tttcttaaag ataatagaga aaaaattgaa 4261 aaaatactta catttagaat accatattat gtaggaccat tagcaagagg aaatagtaga 4321 tttgcttgga tgacaagaaa aagtgaagaa acaataacac catggaattt tgaagaagta 4381 gtagataaag gagcatctgc acaaagtttt atagaaagaa tgacaaattt tgataaaaat 4441 cttccaaatg aaaaagtatt accaaaacat agtttattat atgaatattt tacagtatat 4501 aatgaattaa caaaagtaaa atatgtaaca gaaggtatga gaaaaccagc atttttaagt 4561 ggtgaacaaa aaaaagctat agtagattta ttatttaaaa caaatagaaa agtaacagta 4621 aaacaattaa aagaagatta ttttaaaaaa attgaatgtt ttgatagtgt agaaatatct 4681 ggtgtagaag atagatttaa tgcaagttta ggaacatatc atgatttact taaaataata 4741 aaggataaag attttttaga taatgaagaa aatgaagata ttttagaaga tatagtatta 4801 acattaactt tatttgaaga tagagaaatg atagaagaaa gattaaaaac ttatgctcat 4861 ttatttgatg ataaagtaat gaaacaactt aaaagaagaa gatatacagg ttggggaaga 4921 ttaagtagaa aacttataaa tggaataaga gataaacaaa gtggaaaaac aatattagat 4981 tttcttaaaa gtgatggatt tgcaaataga aattttatgc aattaataca tgatgatagt 5041 ttaacattta aagaagatat tcaaaaagca caagtaagtg gacaaggtga tagtttacat 5101 gaacatatag caaatttagc aggatctcca gcaataaaaa aaggtatatt acaaacagta 5161 aaagtagttg atgaattagt aaaagtaatg ggtagacata aaccagaaaa tatagttata 5221 gaaatggcaa gagaaaatca aacaacacaa aaaggacaaa aaaatagtag agaaagaatg 5281 aaaagaatag aagaaggtat aaaagaatta ggaagtcaaa tacttaaaga acatccagta 5341 gaaaatacac aacttcaaaa tgaaaaactt tatttatatt atttacaaaa tggaagagat 5401 atgtatgtag atcaagaatt agatataaat agattaagtg attatgatgt agatcatata 5461 gtaccacaaa gttttcttaa agatgattct atagataata aagtattaac aagatctgat 5521 aaaaatagag gtaaaagtga taatgtacca agtgaagaag ttgtaaaaaa aatgaaaaat 5581 tattggagac aattacttaa tgcaaaactt ataacacaaa gaaaatttga taatcttaca 5641 aaagcagaaa gaggtggatt atctgaatta gataaagctg gatttataaa aagacaatta 5701 gtagaaacta gacaaataac aaaacatgta gcacaaatat tagatagtag aatgaataca 5761 aaatatgatg aaaatgataa gcttataaga gaagtaaaag ttataacatt aaaaagtaaa 5821 ttagtaagtg attttagaaa agattttcaa ttttataaag taagagaaat aaataattat 5881 catcatgcac atgatgcata tcttaatgca gtagtaggaa cagcattaat aaaaaaatat 5941 cctaaattag aatctgaatt tgtatatgga gattataaag tatatgatgt aagaaaaatg 6001 atagcaaaaa gtgaacaaga aataggaaaa gcaacagcaa agtacttttt ttattctaat 6061 ataatgaatt tttttaagac agaaataact ttagcaaatg gtgaaataag aaaaagacct 6121 ttaatagaaa ctaatggtga aacaggtgaa atagtatggg ataaaggtag agattttgca 6181 acagtaagaa aagtattaag tatgccacaa gtaaatatag taaaaaaaac agaagtacaa 6241 actggaggat ttagtaaaga aagtatactt ccaaaaagaa attcagataa acttattgca 6301 agaaaaaaag attgggatcc aaaaaaatat ggtggatttg atagtccaac agtagcatat 6361 agtgtattag tagtagcaaa agtagaaaaa ggtaaatcta aaaaacttaa aagtgtaaaa 6421 gaattacttg gaataacaat aatggaaaga agtagttttg aaaaaaatcc tattgatttt 6481 ttagaagcaa aaggatataa agaagttaaa aaagatttaa taataaaact tccaaaatat 6541 tctttatttg aattagaaaa tggaagaaaa agaatgttag caagtgctgg tgaattacaa 6601 aaaggtaatg aattagcatt accatctaaa tatgtaaatt ttctttattt agcaagtcat 6661 tatgaaaaac ttaaaggatc tcctgaagat aatgaacaaa aacaattatt tgtagaacaa 6721 cataaacatt atttagatga aataattgaa caaataagtg aattttctaa aagagtaata 6781 ttagcagacg caaatttaga taaagtatta tctgcttata ataaacatag agataaacca 6841 ataagagaac aagcagaaaa tataatacat ctttttacat taacaaattt aggtgcacca 6901 gcagcattta aatattttga tacaacaata gatagaaaaa gatatacaag tacaaaagaa 6961 gtattagacg caacattaat acatcaaagt ataacaggat tatatgaaac aagaatagat 7021 ttatctcaat taggtggtga ttaaaagctt ggcactggcc gtcgttttac aacgtcgtga 7081 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag 7141 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa 7201 tggcgaatgg cgctagcata aaaataagaa gcctgcattt gcaggcttct tatttttatg 7261 gcgcgcccgc ccttaagtct aaaaattagg ggagatgtaa ggatttggga aaaatagaag 7321 atgttataat cataaatatg gtattcgtag gcttaaagtc aaaaaggagg tgaaatataa 7381 atagattttt agctaaatta agtaagaaat aggaggagat ttattgaaca aaaaattaga 7441 aaaaccattt gtatataaga gagagtacga tttgactgga tatgatgttg aaattttaca 7501 aaaatatgag ttagaacaag caatatatgt ttatgttggg agtagttgtg catataacat 7561 gagagctaga agtagtaaat ggagatacca tataagaaca aataataagt ctatatgttg 7621 taacattaaa aattttatac ataacttgga attgttttat aaaatggaat taaagttgtc 7681 agataatatt attaatgata agctatacta tagcaatata gcagagtttg aagaatttga 7741 aacactagaa aaagctagag aggtagaaag tactattata agtcaatatc aatttttaga 7801 ttctataaat cacatgttaa aacaaaaaat aattttattg agtaataagg atagtgtgtt 7861 aaacataact aaaaatggaa atacaaatta tttgaaagta aaaaataaat acatagaaaa 7921 acataagaac aagccaataa tgagatacca tatcaactgt caattcaata cagatggaag 7981 tgtcaaaagt attacacagg agtttgaacc aatattggaa ttaaacaaaa aaaataccct 8041 aagccgacca agcagagtat ttttaaaata atattttaag ataacaacaa aatgagataa 8101 tactactaga caatgacaac tcaactacca attgagttta tggagctacc aactccaata 8161 tcggtctaac tgattaagta tctgtagtta tataataata ttgctatcaa ttttagcatc 8221 ttaacaatat tattatacat actaagctaa aattattcaa tagttgtaaa agttgattag 8281 tcaataagta tatatttaat gtagtgttat ctcttaaaaa aactagataa ggagataata 8341 aatatatgga acaattagat tcaaaatata agttgaaaaa atttctaatg gcagtattta 8401 gagatggtat aggacaagga aataatctta ttgataatga atatgttaga gtatttcaaa 8461 ataataaaag taatagtaaa caattagaac tcggagaaga atttaaagaa tatagtaaaa 8521 caactttttt taaaaatata gatgatatag tagaatttac cttcgcaaaa aatatttatt 8581 atgaaaatac attttttaac ctatgtacta ctgatggaaa agcaggaacc aatgaaaact 8641 taataaatag atatgcatta ggatttgatt ttgacaaaaa agaattagga caaggtttta 8701 attataaaga tataattaat ttatttacta agataggatt acattatcat atcctagttg 8761 atagtggaaa tggattccat gtttatgtgc taattaataa aactaataac attaagttag 8821 tatcagaagt tacaaataca ttaataaata aattgggtgc agataaacaa gcaaatttat 8881 ctactcaagt attaagagta ccttatacat ataatattaa aaatactact aaacaagtaa 8941 aaataataca ccaagacaaa aatatatata gatatgacat agaaaagtta gctaaaaaat 9001 attgcaaaga tgtaaaaaca gtaggtaata ctaatacaaa atatatatta gatagtaagc 9061 taccaaattg tatagtagat attttaaaaa atggtagtaa agatggacat aaaaacctag 9121 atttgcaaaa aatagttgtg actttaagat tgaggaataa aagtttaagt caagtaatat 9181 ccgttgctag agaatggaac tatatatcac aaaatagtct ttcaaatagt gagctagaat 9241 atcaagtcaa gtatatgtat gagaaactta aaacggttaa ttttggttgt actggttgtg 9301 agtttaatag tgattgttgg aataaaatag aatcagattt tatatatagt gatgaagata 9361 ctttgttcaa tatgccacat aagcactcaa aggatttgaa atataagaat aggaaagggg 9421 ttaaaataat gactggtaat caattgttta tctataatgt gttacttaac aataaagata 9481 gagaattaaa catagacgat ataatggagc tgataaccta taaacgtaag aagaaagtta 9541 aaaacattgt tatgagtgaa aagacattaa gagaaacatt aaaagaactt caacataatg 9601 attatattac aaaaacaaaa ggtgttacaa agctaggaat aaaagataca tacaatgtaa 9661 aagaagttag atgtaatata gataaacaat atactattag ttactttgtt accatggcag 9721 taatttgggg aataatttca actgaagaat taagattata tactcacatg agatataagc 9781 aagatttatt ggtcaaagat gataaaataa aaggaaatat attaagaatt aatcaagagg 9841 aattagcaaa agatttagga gtaacacagc aaagaatttc aaatatgata gaatctttat 9901 tagatactaa aattttagat gtatgggaaa ctaaaataaa tgatagagga tttatgtact 9961 atacatatag attaaacaag tagatttttg ataggattag aattgatttt ctagtcctat 10021 ttttatgcaa aaaaactaat taataaaaat ttcttttggt aaaataattg tacgagaatt 10081 gcaaaaaaaa aatggcatca aagtattgaa attaagccgt tttaaaaatt tcttttggta 10141 aaataattct acatatatat gtagtatata tatatatgtt ttttagagaa tgtataacta 10201 gaatatagag ctagaatata gagaatgtat aactagaata tagagctaga atatagagaa 10261 tgtataacta gaatatagag ctagaatata gagaatgtat aactagaata tagagctaga 10321 atatagagaa tgtataacta gaatatagag ctagaatata gagaatgtat aactagaata 10381 tagagctaga atatagagaa tgtataacta gaatatagag ctagaatcct aattagtagg 10441 tgctttttta aaacaagtta aaaatcaaaa atagtattag taagcattgg aaatgctaga 10501 ttctaaaata gaaaagtaaa aaattggtgc actatctaaa cttatctata tcgctttttc 10561 cgtcgtttgg ttctctagtt acgatacagg ggatatgctt atattgagtt atagtactaa 10621 tcagtgctta atatagttaa taaaattata gttaccatag tttagtaact atgatgtatg 10681 ttagttagaa acttgcattt cggccggcca gtgggcaagt tgaaaaattc acaaaaatgt 10741 ggtataatat ctttgttcat tagagcgata aacttgaatt tgagagggaa cttagatggt 10801 atttgaaaaa attgataaaa atagttggaa cagaaaagag tattttgacc actactttgc 10861 aagtgtacct tgtacctaca gcatgaccgt taaagtggat atcacacaaa taaaggaaaa 10921 gggaatgaaa ctatatcctg caatgcttta ttatattgca atgattgtaa accgccattc 10981 agagtttagg acggcaatca atcaagatgg tgaattgggg atatatgatg agatgatacc 11041 aagctataca atatttcaca atgatactga aacattttcc agcctttgga ctgagtgtaa 11101 gtctgacttt aaatcatttt tagcagatta tgaaagtgat acgcaacggt atggaaacaa 11161 tcatagaatg gaaggaaagc caaatgctcc ggaaaacatt tttaatgtat ctatgatacc 11221 gtggtcaacc ttcgatggct ttaatctgaa tttgcagaaa ggatatgatt atttgattcc 11281 tatttttact atggggaaat attataaaga agataacaaa attatacttc ctttggcaat 11341 tcaagttcat cacgcagtat gtgacggatt tcacatttgc cgttttgtaa acgaattgca 11401 ggaattgata aatagttaac ttcaggtttg tctgtaacta aaaacaagta tttaagcaaa 11461 aacatcgtag aaatacggtg ttttttgtta ccctaagttt cataaaaata agaagcctgc 11521 aaatgcaggc ttcttatttt tatggtttaa acggttttag ctgggatatc ggcaaaacaa 11581 gcaaatacta ctatagaagc agttaaaagg gctatttttt ataatctgga gcatagacaa 11641 gaagttagag aagcaattaa aaaagaaaat ataagaaaaa ttcttattat tgggacatca 11701 aaaaaaatgg ttaatcaaat tgtagagaga ttgtcaattg gaagggtctg taagtttata 11761 aatatacaag atataagtac taaatatgaa atagaaatag caaaacaagc tagaaaagaa 11821 ggaaatcatg taattcctgt tccagccgtt gagataaaaa gtatggcaag tggtctcagt 11881 ataaattctt tgaagagact ttttagaaaa ggaaataata ggaatatggt agttttagaa 11941 aaaacaatta taagaccaac ttttagttat ataggaaagt tttatattag tgctgatgta 12001 attaaacaga ttattgaata tgaagtatac aactttggaa gcattgatag aataaataaa 12061 atcaatatag agaattgcaa taattttatg aatatttttg taagtataaa tatcaatgat 12121 ttagaaacta taaaaaaaat tgaagacata caacagacaa taaaaaagtc tattgagaaa 12181 atgactttag ttaatgtaca aaaaatagat attaacattc ataaaattaa aaaaataaac 12241 ttcctccaat gttaaaaaat aaactgagaa aatgatatac taatttgcaa taaagtgtgc 12301 tataattaaa ctgtaaaggt accgaaaagt gatgcattgt tgggttttag agctagaaat 12361 agcaagttaa aataaggcta gtccgttatc aacttgaaaa agtggcaccg agtcggtgct 12421 ttttttctat ggagaaatct agatcagcat gatgtctgac tagacgcgta agctctgcaa 12481 ctatttttag atggttgcag agcttatttt tattttttat taaactcctt tttgataatc 12541 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa 12601 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa 12661 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc 12721 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt 12781 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc 12841 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac 12901 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca 12961 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg 13021 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag 13081 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt 13141 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat 13201 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc 13261 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt 13321 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag 13381 cggaagagcg cccaatacgc agggccctaa catcttctat ttttcccaaa tccttacatc 13441 tcccctaatt tttagactta agggcgaatt cgcccttttt tatactcccc ttgtaccaaa 13501 aggatatgat ttcttggagg aaattaaaaa gttatccaac agcagacagt ccacgcaggc 13561 gacgtgcgaa gcggaagtcg caggtgtgga ctgatcttgc tggctggtgt ggcaatagcc 13621 acgccagcac ttaacccccc gtatctaaca ggggggtaca aatcgacagg aaacagtcaa 13681 aaaaacatta gaaaatcctt tggttacaag ggatttacaa aatttcagcg tatgtcaaat 13741 gggctttaaa agttgacata cggccttttt gattggaggg atttttactg aatgaacaaa 13801 cttggttaca gcatttaaaa gaaaaacgct tggcttatgg actatctcaa aaccgtttag 13861 ctgttgcgac tggtattaca aggcagtatc taagcgatat tgaaacagga aaagtcaagc 13921 catcagagga tttacagcag tccctttggg aagctctgga acgcttcaat cccgacgctc 13981 cccttgaaat gctgtttgat tatgtaagga ttcgctttcc gacaacagac gtacagcagg 14041 tggtcgaaaa catcttacaa ctgaaactgt cctattttct tcatgaggac tatggtttct 14101 attcttattc agagcattat gctttaggcg acatattcgt cctttgaggg cgaattctgc 14161 agattaccta ataatttatc tacattccct ttaggggccc //