LOCUS Exported 16437 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION CRISPR/Cas based plant genome editing and gene regulation; expresses 3×FLAG-NLS-zCas9-NLS, gRNA scaffold for insertion of target sequence (AtU6-26 promoter), Kan resistance. ACCESSION . VERSION . KEYWORDS pKSE401 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 16437) AUTHORS Xing HL, Dong L, Wang ZP, Zhang HY, Han CY, Liu B, Wang XC, Chen QJ TITLE A CRISPR/Cas9 toolkit for multiplex genome editing in plants. JOURNAL BMC Plant Biol. 2014 Nov 29;14(1):327. PUBMED 25432517 REFERENCE 2 (bases 1 to 16437) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..16437 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 21..45 /label=RB T-DNA repeat /note="right border repeat from nopaline C58 T-DNA" primer_bind 233..255 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 247..264 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 248..264 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" CDS 977..1768 /codon_start=1 /gene="aadA" /product="aminoglycoside adenylyltransferase (Murphy, 1985)" /label=SmR /note="confers resistance to spectinomycin and streptomycin" /translation="MGEAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK" misc_RNA 1920..1995 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter 2609..2953 /label=CaMV 35S promoter /note="strong constitutive promoter from cauliflower mosaic virus" CDS 3106..3171 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS 3178..3198 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 3223..7323 /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD ATLIHQSITGLYETRIDLSQLGGD" RBS 7236..7244 /label=Shine-Dalgarno sequence /note="full consensus sequence for ribosome-binding sites upstream of start codons in E. coli; complementary to a region in the 3' end of the 16S rRNA (Chen et al., 1994)" CDS 7324..7371 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" primer_bind complement(8028..8044) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(8028..8044) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(8041..8063) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 8052..8068 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." promoter complement(8076..8106) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 8121..8142 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 8333..9009 /label=CaMV 35S promoter (enhanced) /note="cauliflower mosaic virus 35S promoter with a duplicated enhancer region" CDS 9072..9869 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MGIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLL SSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEH QGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDI ALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(9129..9148) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 9739..9758 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 9926..10100 /label=CaMV poly(A) signal /note="cauliflower mosaic virus polyadenylation signal" misc_feature 10178..10202 /label=LB T-DNA repeat /note="left border repeat from nopaline C58 T-DNA" CDS 10627..11421 /codon_start=1 /gene="aphA-3" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin" /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF" rep_origin 11508..12096 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 11997..12016 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 12250..12267 /label=L4440 /note="L4440 vector, forward primer" misc_feature 12282..12422 /label=bom /note="basis of mobility region from pBR322" primer_bind complement(12348..12367) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 12508..12530 /label=pGEX 3' /note="pGEX vectors, reverse primer" rep_origin 12766..12960 /label=pVS1 oriV /note="origin of replication for the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" CDS complement(13026..14099) /codon_start=1 /product="replication protein from the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" /label=pVS1 RepA /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI LVMRYRNLIEGEASAGS" CDS complement(14528..15157) /codon_start=1 /product="stability protein from the Pseudomonas plasmid pVS1 (Heeb et al., 2000)" /label=pVS1 StaA /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI" ORIGIN 1 taaacgctct tttctcttag gtttacccgc caatatatcc tgtcaaacac tgatagttta 61 aactgaaggc gggaaacgac aatctgatcc aagctcaagc tgctctagca ttcgccattc 121 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg 181 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca 241 cgacgttgta aaacgacggc cagtgccaag cttcgacttg ccttccgcac aatacatcat 301 ttcttcttag ctttttttct tcttcttcgt tcatacagtt tttttttgtt tatcagctta 361 cattttcttg aaccgtagct ttcgttttct tctttttaac tttccattcg gagtttttgt 421 atcttgtttc atagtttgtc ccaggattag aatgattagg catcgaacct tcaagaattt 481 gattgaataa aacatcttca ttcttaagat atgaagataa tcttcaaaag gcccctggga 541 atctgaaaga agagaagcag gcccatttat atgggaaaga acaatagtat ttcttatata 601 ggcccattta agttgaaaac aatcttcaaa agtcccacat cgcttagata agaaaacgaa 661 gctgagttta tatacagcta gagtcgaagt agtgattggg agaccaaccc agtggacata 721 agcctgttcg gttcgtaagc tgtaatgcaa gtagcgtatg cgctcacgca actggtccag 781 aaccttgacc gaacgcagcg gtggtaacgg cgcagtggcg gttttcatgg cttgttatga 841 ctgttttttt ggggtacagt ctatgcctcg ggcatccaag cagcaagcgc gttacgccgt 901 gggtcgatgt ttgatgttat ggagcagcaa cgatgttacg cagcagggca gtcgccctaa 961 aacaaagtta aacatcatgg gggaagcggt gatcgccgaa gtatcgactc aactatcaga 1021 ggtagttggc gtcatcgagc gccatctcga accgacgttg ctggccgtac atttgtacgg 1081 ctccgcagtg gatggcggcc tgaagccaca cagtgatatt gatttgctgg ttacggtgac 1141 cgtaaggctt gatgaaacaa cgcggcgagc tttgatcaac gaccttttgg aaacttcggc 1201 ttcccctgga gagagcgaga ttctccgcgc tgtagaagtc accattgttg tgcacgacga 1261 catcattccg tggcgttatc cagctaagcg cgaactgcaa tttggagaat ggcagcgcaa 1321 tgacattctt gcaggtatct tcgagccagc cacgatcgac attgatctgg ctatcttgct 1381 gacaaaagca agagaacata gcgttgcctt ggtaggtcca gcggcggagg aactctttga 1441 tccggttcct gaacaggatc tatttgaggc gctaaatgaa accttaacgc tatggaactc 1501 gccgcccgac tgggctggcg atgagcgaaa tgtagtgctt acgttgtccc gcatttggta 1561 cagcgcagta accggcaaaa tcgcgccgaa ggatgtcgct gccgactggg caatggagcg 1621 cctgccggcc cagtatcagc ccgtcatact tgaagctaga caggcttatc ttggacaaga 1681 agaagatcgc ttggcctcgc gcgcagatca gttggaagaa tttgtccact acgtgaaagg 1741 cgagatcacc aaggtagtcg gcaaataatg tctagctaga aattcgttca agccgacgcc 1801 gcttcgcggc gcggcttaac tcaagcgtta gatgcactaa gcacataatt gctcacagcc 1861 aaactatcag gtcaagtctg cttttattat ttttaagcgt gcataataag ccggtctcgg 1921 ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg 1981 gcaccgagtc ggtgcttttt tttgcaaaat tttccagatc gatttcttct tcctctgttc 2041 ttcggcgttc aatttctggg gttttctctt cgttttctgt aactgaaacc taaaatttga 2101 cctaaaaaaa atctcaaata atatgattca gtggttttgt acttttcagt tagttgagtt 2161 ttgcagttcc gatgagataa accaataagc ttgcatgcct gcaggtcaac atggtggagc 2221 acgacacact tgtctactcc aaaaatatca aagatacagt ctcagaagac caaagggcaa 2281 ttgagacttt tcaacaaagg gtaatatccg gaaacctcct cggattccat tgcccagcta 2341 tctgtcactt tattgtgaag atagtggaaa aggaaggtgg ctcctacaaa tgccatcatt 2401 gcgataaagg aaaggccatc gttgaagatg cctctgccga cagtggtccc aaagatggac 2461 ccccacccac gaggagcatc gtggaaaaag aagacgttcc aaccacgtct tcaaagcaag 2521 tggattgatg tgataacatg gtggagcacg acacacttgt ctactccaaa aatatcaaag 2581 atacagtctc agaagaccaa agggcaattg agacttttca acaaagggta atatccggaa 2641 acctcctcgg attccattgc ccagctatct gtcactttat tgtgaagata gtggaaaagg 2701 aaggtggctc ctacaaatgc catcattgcg ataaaggaaa ggccatcgtt gaagatgcct 2761 ctgccgacag tggtcccaaa gatggacccc cacccacgag gagcatcgtg gaaaaagaag 2821 acgttccaac cacgtcttca aagcaagtgg attgatgtga tatctccact gacgtaaggg 2881 atgacgcaca atcccactat ccttcgcaag acccttcctc tatataagga agttcatttc 2941 atttggagag gacctcgacc tcaacacaac atatacaaaa caaacgaatc tcaagcaatc 3001 aagcattcta cttctattgc agcaatttaa atcatttctt ttaaagcaaa agcaattttc 3061 tgaaaatttt caccatttac gaacgatact cgagtaatct agatggatta caaggaccac 3121 gacggggatt acaaggacca cgacattgat tacaaggatg atgatgacaa gatggctccg 3181 aagaagaaga ggaaggttgg catccacggg gtgccagctg ctgacaagaa gtactcgatc 3241 ggcctcgata ttgggactaa ctctgttggc tgggccgtga tcaccgacga gtacaaggtg 3301 ccctcaaaga agttcaaggt cctgggcaac accgatcggc attccatcaa gaagaatctc 3361 attggcgctc tcctgttcga cagcggcgag acggctgagg ctacgcggct caagcgcacc 3421 gcccgcaggc ggtacacgcg caggaagaat cgcatctgct acctgcagga gattttctcc 3481 aacgagatgg cgaaggttga cgattctttc ttccacaggc tggaggagtc attcctcgtg 3541 gaggaggata agaagcacga gcggcatcca atcttcggca acattgtcga cgaggttgcc 3601 taccacgaga agtaccctac gatctaccat ctgcggaaga agctcgtgga ctccacagat 3661 aaggcggacc tccgcctgat ctacctcgct ctggcccaca tgattaagtt caggggccat 3721 ttcctgatcg agggggatct caacccggac aatagcgatg ttgacaagct gttcatccag 3781 ctcgtgcaga cgtacaacca gctcttcgag gagaacccca ttaatgcgtc aggcgtcgac 3841 gcgaaggcta tcctgtccgc taggctctcg aagtctcggc gcctcgagaa cctgatcgcc 3901 cagctgccgg gcgagaagaa gaacggcctg ttcgggaatc tcattgcgct cagcctgggg 3961 ctcacgccca acttcaagtc gaatttcgat ctcgctgagg acgccaagct gcagctctcc 4021 aaggacacat acgacgatga cctggataac ctcctggccc agatcggcga tcagtacgcg 4081 gacctgttcc tcgctgccaa gaatctgtcg gacgccatcc tcctgtctga tattctcagg 4141 gtgaacaccg agattacgaa ggctccgctc tcagcctcca tgatcaagcg ctacgacgag 4201 caccatcagg atctgaccct cctgaaggcg ctggtcaggc agcagctccc cgagaagtac 4261 aaggagatct tcttcgatca gtcgaagaac ggctacgctg ggtacattga cggcggggcc 4321 tctcaggagg agttctacaa gttcatcaag ccgattctgg agaagatgga cggcacggag 4381 gagctgctgg tgaagctcaa tcgcgaggac ctcctgagga agcagcggac attcgataac 4441 ggcagcatcc cacaccagat tcatctcggg gagctgcacg ctatcctgag gaggcaggag 4501 gacttctacc ctttcctcaa ggataaccgc gagaagatcg agaagattct gactttcagg 4561 atcccgtact acgtcggccc actcgctagg ggcaactccc gcttcgcttg gatgacccgc 4621 aagtcagagg agacgatcac gccgtggaac ttcgaggagg tggtcgacaa gggcgctagc 4681 gctcagtcgt tcatcgagag gatgacgaat ttcgacaaga acctgccaaa tgagaaggtg 4741 ctccctaagc actcgctcct gtacgagtac ttcacagtct acaacgagct gactaaggtg 4801 aagtatgtga ccgagggcat gaggaagccg gctttcctgt ctggggagca gaagaaggcc 4861 atcgtggacc tcctgttcaa gaccaaccgg aaggtcacgg ttaagcagct caaggaggac 4921 tacttcaaga agattgagtg cttcgattcg gtcgagatct ctggcgttga ggaccgcttc 4981 aacgcctccc tggggaccta ccacgatctc ctgaagatca ttaaggataa ggacttcctg 5041 gacaacgagg agaatgagga tatcctcgag gacattgtgc tgacactcac tctgttcgag 5101 gaccgggaga tgatcgagga gcgcctgaag acttacgccc atctcttcga tgacaaggtc 5161 atgaagcagc tcaagaggag gaggtacacc ggctggggga ggctgagcag gaagctcatc 5221 aacggcattc gggacaagca gtccgggaag acgatcctcg acttcctgaa gagcgatggc 5281 ttcgcgaacc gcaatttcat gcagctgatt cacgatgaca gcctcacatt caaggaggat 5341 atccagaagg ctcaggtgag cggccagggg gactcgctgc acgagcatat cgcgaacctc 5401 gctggctcgc cagctatcaa gaaggggatt ctgcagaccg tgaaggttgt ggacgagctg 5461 gtgaaggtca tgggcaggca caagcctgag aacatcgtca ttgagatggc ccgggagaat 5521 cagaccacgc agaagggcca gaagaactca cgcgagagga tgaagaggat cgaggagggc 5581 attaaggagc tggggtccca gatcctcaag gagcacccgg tggagaacac gcagctgcag 5641 aatgagaagc tctacctgta ctacctccag aatggccgcg atatgtatgt ggaccaggag 5701 ctggatatta acaggctcag cgattacgac gtcgatcata tcgttccaca gtcattcctg 5761 aaggatgact ccattgacaa caaggtcctc accaggtcgg acaagaaccg gggcaagtct 5821 gataatgttc cttcagagga ggtcgttaag aagatgaaga actactggcg ccagctcctg 5881 aatgccaagc tgatcacgca gcggaagttc gataacctca caaaggctga gaggggcggg 5941 ctctctgagc tggacaaggc gggcttcatc aagaggcagc tggtcgagac acggcagatc 6001 actaagcacg ttgcgcagat tctcgactca cggatgaaca ctaagtacga tgagaatgac 6061 aagctgatcc gcgaggtgaa ggtcatcacc ctgaagtcaa agctcgtctc cgacttcagg 6121 aaggatttcc agttctacaa ggttcgggag atcaacaatt accaccatgc ccatgacgcg 6181 tacctgaacg cggtggtcgg cacagctctg atcaagaagt acccaaagct cgagagcgag 6241 ttcgtgtacg gggactacaa ggtttacgat gtgaggaaga tgatcgccaa gtcggagcag 6301 gagattggca aggctaccgc caagtacttc ttctactcta acattatgaa tttcttcaag 6361 acagagatca ctctggccaa tggcgagatc cggaagcgcc ccctcatcga gacgaacggc 6421 gagacggggg agatcgtgtg ggacaagggc agggatttcg cgaccgtcag gaaggttctc 6481 tccatgccac aagtgaatat cgtcaagaag acagaggtcc agactggcgg gttctctaag 6541 gagtcaattc tgcctaagcg gaacagcgac aagctcatcg cccgcaagaa ggactgggat 6601 ccgaagaagt acggcgggtt cgacagcccc actgtggcct actcggtcct ggttgtggcg 6661 aaggttgaga agggcaagtc caagaagctc aagagcgtga aggagctgct ggggatcacg 6721 attatggagc gctccagctt cgagaagaac ccgatcgatt tcctggaggc gaagggctac 6781 aaggaggtga agaaggacct gatcattaag ctccccaagt actcactctt cgagctggag 6841 aacggcagga agcggatgct ggcttccgct ggcgagctgc agaaggggaa cgagctggct 6901 ctgccgtcca agtatgtgaa cttcctctac ctggcctccc actacgagaa gctcaagggc 6961 agccccgagg acaacgagca gaagcagctg ttcgtcgagc agcacaagca ttacctcgac 7021 gagatcattg agcagatttc cgagttctcc aagcgcgtga tcctggccga cgcgaatctg 7081 gataaggtcc tctccgcgta caacaagcac cgcgacaagc caatcaggga gcaggctgag 7141 aatatcattc atctcttcac cctgacgaac ctcggcgccc ctgctgcttt caagtacttc 7201 gacacaacta tcgatcgcaa gaggtacaca agcactaagg aggtcctgga cgcgaccctc 7261 atccaccagt cgattaccgg cctctacgag acgcgcatcg acctgtctca gctcgggggc 7321 gacaagcggc cagcggcgac gaagaaggcg gggcaggcga agaagaagaa gtgagctcag 7381 agctttcgtt cgtatcatcg gtttcgacaa cgttcgtcaa gttcaatgca tcagtttcat 7441 tgcgcacaca ccagaatcct actgagtttg agtattatgg cattgggaaa actgtttttc 7501 ttgtaccatt tgttgtgctt gtaatttact gtgtttttta ttcggttttc gctatcgaac 7561 tgtgaaatgg aaatggatgg agaagagtta atgaatgata tggtcctttt gttcattctc 7621 aaattaatat tatttgtttt ttctcttatt tgttgtgtgt tgaatttgaa attataagag 7681 atatgcaaac attttgtttt gagtaaaaat gtgtcaaatc gtggcctcta atgaccgaag 7741 ttaatatgag gagtaaaaca cttgtagttg taccattatg cttattcact aggcaacaaa 7801 tatattttca gacctagaaa agctgcaaat gttactgaat acaagtatgt cctcttgtgt 7861 tttagacatt tatgaacttt cctttatgta attttccaga atccttgtca gattctaatc 7921 attgctttat aattatagtt atactcatgg atttgtagtt gagtatgaaa atatttttta 7981 atgcatttta tgacttgcca attgattgac aacgaattcg taatcatgtc atagctgttt 8041 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag 8101 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg 8161 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg 8221 gggagaggcg gtttgcgtat tggctagagc agcttgccaa catggtggag cacgacactc 8281 tcgtctactc caagaatatc aaagatacag tctcagaaga ccaaagggct attgagactt 8341 ttcaacaaag ggtaatatcg ggaaacctcc tcggattcca ttgcccagct atctgtcact 8401 tcatcaaaag gacagtagaa aaggaaggtg gcacctacaa atgccatcat tgcgataaag 8461 gaaaggctat cgttcaagat gcctctgccg acagtggtcc caaagatgga cccccaccca 8521 cgaggagcat cgtggaaaaa gaagacgttc caaccacgtc ttcaaagcaa gtggattgat 8581 gtgaacatgg tggagcacga cactctcgtc tactccaaga atatcaaaga tacagtctca 8641 gaagaccaaa gggctattga gacttttcaa caaagggtaa tatcgggaaa cctcctcgga 8701 ttccattgcc cagctatctg tcacttcatc aaaaggacag tagaaaagga aggtggcacc 8761 tacaaatgcc atcattgcga taaaggaaag gctatcgttc aagatgcctc tgccgacagt 8821 ggtcccaaag atggaccccc acccacgagg agcatcgtgg aaaaagaaga cgttccaacc 8881 acgtcttcaa agcaagtgga ttgatgtgat atctccactg acgtaaggga tgacgcacaa 8941 tcccactatc cttcgcaaga cccttcctct atataaggaa gttcatttca tttggagagg 9001 acacgctgaa atcaccagtc tctctctaca aatctatctc tctcgagctt tcgcagatct 9061 gtcgatcgac catggggatt gaacaagatg gattgcacgc aggttctccg gccgcttggg 9121 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg 9181 tgtttcggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg 9241 ccctgaatga actccaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc 9301 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg 9361 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca 9421 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc 9481 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg 9541 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg 9601 cgcgcatgcc cgacggcgag gatctcgtcg tgacacatgg cgatgcctgc ttgccgaata 9661 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg 9721 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat 9781 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct 9841 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcggat cgatcctcta 9901 gctagagtcg atcgacaagc tcgagtttct ccataataat gtgtgagtag ttcccagata 9961 agggaattag ggttcctata gggtttcgct catgtgttga gcatataaga aacccttagt 10021 atgtatttgt atttgtaaaa tacttctatc aataaaattt ctaattccta aaaccaaaat 10081 ccagtactaa aatccagatc ccccgaatta attcggcgtt aattcagtac attaaaaacg 10141 tccgcaatgt gttattaagt tgtctaagcg tcaatttgtt tacaccacaa tatatcctgc 10201 caccagccag ccaacagctc cccgaccggc agctcggcac aaaatcacca ctcgatacag 10261 gcagcccatc agtccgggac ggcgtcagcg ggagagccgt tgtaaggcgg cagactttgc 10321 tcatgttacc gatgctattc ggaagaacgg caactaagct gccgggtttg aaacacggat 10381 gatctcgcgg agggtagcat gttgattgta acgatgacag agcgttgctg cctgtgatca 10441 ccgcggtttc aaaatcggct ccgtcgatac tatgttatac gccaactttg aaaacaactt 10501 tgaaaaagct gttttctggt atttaaggtt ttagaatgca aggaacagtg aattggagtt 10561 cgtcttgtta taattagctt cttggggtat ctttaaatac tgtagaaaag aggaaggaaa 10621 taataaatgg ctaaaatgag aatatcaccg gaattgaaaa aactgatcga aaaataccgc 10681 tgcgtaaaag atacggaagg aatgtctcct gctaaggtat ataagctggt gggagaaaat 10741 gaaaacctat atttaaaaat gacggacagc cggtataaag ggaccaccta tgatgtggaa 10801 cgggaaaagg acatgatgct atggctggaa ggaaagctgc ctgttccaaa ggtcctgcac 10861 tttgaacggc atgatggctg gagcaatctg ctcatgagtg aggccgatgg cgtcctttgc 10921 tcggaagagt atgaagatga acaaagccct gaaaagatta tcgagctgta tgcggagtgc 10981 atcaggctct ttcactccat cgacatatcg gattgtccct atacgaatag cttagacagc 11041 cgcttagccg aattggatta cttactgaat aacgatctgg ccgatgtgga ttgcgaaaac 11101 tgggaagaag acactccatt taaagatccg cgcgagctgt atgatttttt aaagacggaa 11161 aagcccgaag aggaacttgt cttttcccac ggcgacctgg gagacagcaa catctttgtg 11221 aaagatggca aagtaagtgg ctttattgat cttgggagaa gcggcagggc ggacaagtgg 11281 tatgacattg ccttctgcgt ccggtcgatc agggaggata tcggggaaga acagtatgtc 11341 gagctatttt ttgacttact ggggatcaag cctgattggg agaaaataaa atattatatt 11401 ttactggatg aattgtttta gtacctagaa tgcatgacca aaatccctta acgtgagttt 11461 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt 11521 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt 11581 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag 11641 ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta 11701 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat 11761 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg 11821 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg 11881 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac 11941 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga 12001 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt 12061 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta 12121 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat 12181 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg 12241 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc 12301 cttacgcatc tgtgcggtat ttcacaccgc atatggtgca ctctcagtac aatctgctct 12361 gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg 12421 cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat 12481 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt 12541 catcaccgaa acgcgcgagg cagggtgcct tgatgtgggc gccggcggtc gagtggcgac 12601 ggcgcggctt gtccgcgccc tggtagattg cctggccgta ggccagccat ttttgagcgg 12661 ccagcggccg cgataggccg acgcgaagcg gcggggcgta gggagcgcag cgaccgaagg 12721 gtaggcgctt tttgcagctc ttcggctgtg cgctggccag acagttatgc acaggccagg 12781 cgggttttaa gagttttaat aagttttaaa gagttttagg cggaaaaatc gccttttttc 12841 tcttttatat cagtcactta catgtgtgac cggttcccaa tgtacggctt tgggttccca 12901 atgtacgggt tccggttccc aatgtacggc tttgggttcc caatgtacgt gctatccaca 12961 ggaaacagac cttttcgacc tttttcccct gctagggcaa tttgccctag catctgctcc 13021 gtacattagg aaccggcgga tgcttcgccc tcgatcaggt tgcggtagcg catgactagg 13081 atcgggccag cctgccccgc ctcctccttc aaatcgtact ccggcaggtc atttgacccg 13141 atcagcttgc gcacggtgaa acagaacttc ttgaactctc cggcgctgcc actgcgttcg 13201 tagatcgtct tgaacaacca tctggcttct gccttgcctg cggcgcggcg tgccaggcgg 13261 tagagaaaac ggccgatgcc gggatcgatc aaaaagtaat cggggtgaac cgtcagcacg 13321 tccgggttct tgccttctgt gatctcgcgg tacatccaat cagctagctc gatctcgatg 13381 tactccggcc gcccggtttc gctctttacg atcttgtagc ggctaatcaa ggcttcaccc 13441 tcggataccg tcaccaggcg gccgttcttg gccttcttcg tacgctgcat ggcaacgtgc 13501 gtggtgttta accgaatgca ggtttctacc aggtcgtctt tctgctttcc gccatcggct 13561 cgccggcaga acttgagtac gtccgcaacg tgtggacgga acacgcggcc gggcttgtct 13621 cccttccctt cccggtatcg gttcatggat tcggttagat gggaaaccgc catcagtacc 13681 aggtcgtaat cccacacact ggccatgccg gccggccctg cggaaacctc tacgtgcccg 13741 tctggaagct cgtagcggat cacctcgcca gctcgtcggt cacgcttcga cagacggaaa 13801 acggccacgt ccatgatgct gcgactatcg cgggtgccca cgtcatagag catcggaacg 13861 aaaaaatctg gttgctcgtc gcccttgggc ggcttcctaa tcgacggcgc accggctgcc 13921 ggcggttgcc gggattcttt gcggattcga tcagcggccg cttgccacga ttcaccgggg 13981 cgtgcttctg cctcgatgcg ttgccgctgg gcggcctgcg cggccttcaa cttctccacc 14041 aggtcatcac ccagcgccgc gccgatttgt accgggccgg atggtttgcg accgctcacg 14101 ccgattcctc gggcttgggg gttccagtgc cattgcaggg ccggcaggca acccagccgc 14161 ttacgcctgg ccaaccgccc gttcctccac acatggggca ttccacggcg tcggtgcctg 14221 gttgttcttg attttccatg ccgcctcctt tagccgctaa aattcatcta ctcatttatt 14281 catttgctca tttactctgg tagctgcgcg atgtattcag atagcagctc ggtaatggtc 14341 ttgccttggc gtaccgcgta catcttcagc ttggtgtgat cctccgccgg caactgaaag 14401 ttgacccgct tcatggctgg cgtgtctgcc aggctggcca acgttgcagc cttgctgctg 14461 cgtgcgctcg gacggccggc acttagcgtg tttgtgcttt tgctcatttt ctctttacct 14521 cattaactca aatgagtttt gatttaattt cagcggccag cgcctggacc tcgcgggcag 14581 cgtcgccctc gggttctgat tcaagaacgg ttgtgccggc ggcggcagtg cctgggtagc 14641 tcacgcgctg cgtgatacgg gactcaagaa tgggcagctc gtacccggcc agcgcctcgg 14701 caacctcacc gccgatgcgc gtgcctttga tcgcccgcga cacgacaaag gccgcttgta 14761 gccttccatc cgtgacctca atgcgctgct taaccagctc caccaggtcg gcggtggccc 14821 atatgtcgta agggcttggc tgcaccggaa tcagcacgaa gtcggctgcc ttgatcgcgg 14881 acacagccaa gtccgccgcc tggggcgctc cgtcgatcac tacgaagtcg cgccggccga 14941 tggccttcac gtcgcggtca atcgtcgggc ggtcgatgcc gacaacggtt agcggttgat 15001 cttcccgcac ggccgcccaa tcgcgggcac tgccctgggg atcggaatcg actaacagaa 15061 catcggcccc ggcgagttgc agggcgcggg ctagatgggt tgcgatggtc gtcttgcctg 15121 acccgccttt ctggttaagt acagcgataa ccttcatgcg ttccccttgc gtatttgttt 15181 atttactcat cgcatcatat acgcagcgac cgcatgacgc aagctgtttt actcaaatac 15241 acatcacctt tttagacggc ggcgctcggt ttcttcagcg gccaagctgg ccggccaggc 15301 cgccagcttg gcatcagaca aaccggccag gatttcatgc agccgcacgg ttgagacgtg 15361 cgcgggcggc tcgaacacgt acccggccgc gatcatctcc gcctcgatct cttcggtaat 15421 gaaaaacggt tcgtcctggc cgtcctggtg cggtttcatg cttgttcctc ttggcgttca 15481 ttctcggcgg ccgccagggc gtcggcctcg gtcaatgcgt cctcacggaa ggcaccgcgc 15541 cgcctggcct cggtgggcgt cacttcctcg ctgcgctcaa gtgcgcggta cagggtcgag 15601 cgatgcacgc caagcagtgc agccgcctct ttcacggtgc ggccttcctg gtcgatcagc 15661 tcgcgggcgt gcgcgatctg tgccggggtg agggtagggc gggggccaaa cttcacgcct 15721 cgggccttgg cggcctcgcg cccgctccgg gtgcggtcga tgattaggga acgctcgaac 15781 tcggcaatgc cggcgaacac ggtcaacacc atgcggccgg ccggcgtggt ggtgtcggcc 15841 cacggctctg ccaggctacg caggcccgcg ccggcctcct ggatgcgctc ggcaatgtcc 15901 agtaggtcgc gggtgctgcg ggccaggcgg tctagcctgg tcactgtcac aacgtcgcca 15961 gggcgtaggt ggtcaagcat cctggccagc tccgggcggt cgcgcctggt gccggtgatc 16021 ttctcggaaa acagcttggt gcagccggcc gcgtgcagtt cggcccgttg gttggtcaag 16081 tcctggtcgt cggtgctgac gcgggcatag cccagcaggc cagcggcggc gctcttgttc 16141 atggcgtaat gtctccggtt ctagtcgcaa gtattctact ttatgcgact aaaacacgcg 16201 acaagaaaac gccaggaaaa gggcagggcg gcagcctgtc gcgtaactta ggacttgtgc 16261 gacatgtcgt tttcagaaga cggctgcact gaacgtcaga agccgactgc actatagcag 16321 cggaggggtt ggatcaaagt actttgatcc cgaggggaac cctgtggttg gcatgcacat 16381 acaaatggac gaacggataa accttttcac gcccttttaa atatccgatt attctaa //