LOCUS Exported 12671 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Recombineering plasmid with constitutively expressed cas9 and the araBAD promoter expressing alpha-beta-gamma. ACCESSION . VERSION . KEYWORDS pCas9 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 12671) AUTHORS Cook TB, Rand JM, Nurani W, Courtney DK, Liu SA, Pfleger BF TITLE Genetic tools for reliable gene expression and recombineering in Pseudomonas putida. JOURNAL J Ind Microbiol Biotechnol. 2018 Jan 3. pii: 10.1007/s10295-017-2001-5. doi: 10.1007/s10295-017-2001-5. PUBMED 29299733 REFERENCE 2 (bases 1 to 12671) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..12671 /organism="synthetic DNA construct" /mol_type="other DNA" CDS complement(62..1210) /codon_start=1 /product="trans-acting replication protein that binds to and activates oriV" /label=trfA /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS IVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR NKKIPREALKNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR" rep_origin 1603..2134 /label=oriV /note="incP origin of replication" CDS complement(2549..3082) /codon_start=1 /gene="aacC1" /product="gentamycin acetyltransferase" /label=GmR /note="confers resistance to gentamycin" /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEVVVGALAAYVLPKFEQPRS EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR EEVMHFDIDPSTAT" promoter complement(3271..3299) /gene="intI1 (promoter lies within the coding sequence)" /label=Pc promoter /note="class 1 integron promoter" CDS complement(3514..3537) /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS complement(3547..7650) /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" misc_feature 7721..7742 /label=BioBrick prefix /note="BioBrick prefix for parts that do not start with ""ATG""" terminator 7792..7838 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" primer_bind 7991..8008 /label=pBAD Reverse /note="For vectors with E. coli araBAD promoter, reverse primer" CDS complement(8064..8081) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS complement(8097..8126) /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" misc_RNA complement(8147..8222) /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" protein_bind 8253..8271 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 8278..8296 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" promoter 8353..8430 /gene="lacI" /label=lacI promoter CDS 8431..9054 /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 complement(8504..8523) /label=Tet-R /note="Tetracycline resistance gene, reverse primer" terminator 9188..9219 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" CDS complement(9270..10148) /codon_start=1 /gene="araC" /product="L-arabinose regulatory protein" /label=araC /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE EKVNDVAVKLS" promoter 10175..10459 /gene="araBAD" /label=araBAD promoter /note="promoter of the L-arabinose operon of E. coli; the araC regulatory gene is transcribed in the opposite direction (Guzman et al., 1995)" primer_bind 10382..10401 /label=pBAD Forward /note="For vectors with E. coli araBAD promoter, forward primer" RBS 10486..10494 /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 10503..10919 /codon_start=1 /gene="gam" /product="inhibitor of the host RecBCD nuclease in the lambda Red system" /label=Gam /note="protects the double-stranded DNA substrate for recombination" /translation="MDINTETEIKQKHSLTPFPVFLISPAFRGRYFHSYFRSSAMNAYY IQDRLEAQSWARHYQQLAREEKEAELADDMEKGLPQHLFESLCIDHLQRHGASKKSITR AFDDDVEFQERMAEHIRYMVETIAHHQVDIDSEV" CDS 10925..11710 /codon_start=1 /gene="bet" /product="single-stranded DNA binding recombinase in the lambda Red system" /label=Beta /note="mediates pairing of a single-stranded DNA with a complementary target" /translation="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFI ALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESC TCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLA FGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLP LCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA" CDS 11707..12387 /codon_start=1 /gene="exo" /product="5' to 3' double-stranded DNA exonuclease in the lambda Red system" /label=Exo /note="creates 3' single-stranded DNA tails" /translation="MTPDIILQRTGIDVRAVEQGDDAWHKLRLGVITASEVHNVIAKPR SGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESP IIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMW VTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFG EQWR" terminator 12388..12632 /label=-lambda tL3 terminator /note="transcription terminator tL3 from phage lambda" ORIGIN 1 ttatacctag ggcgatgcag gtggctgctg aacccccagc cggaactgac cccacaaggc 61 cctagcgttt gcaatgcacc aggtcatcat tgacccaggc gtgttccacc aggccgctgc 121 ctcgcaactc ttcgcaggct tcgccgacct gctcgcgcca cttcttcacg cgggtggaat 181 ccgatccgca catgaggcgg aaggtttcca gcttgagcgg gtacggctcc cggtgcgagc 241 tgaaatagtc gaacatccgt cgggccgtcg gcgacagctt gcggtacttc tcccatatga 301 atttcgtgta gtggtcgcca gcaaacagca cgacgatttc ctcgtcgatc aggacctggc 361 aacgggacgt tttcttgcca cggtccagga cgcggaagcg gtgcagcagc gacaccgatt 421 ccaggtgccc aacgcggtcg gacgtgaagc ccatcgccgt cgcctgtagg cgcgacaggc 481 attcctcggc cttcgtgtaa taccggccat tgatcgacca gcccaggtcc tggcaaagct 541 cgtagaacgt gaaggtgatc ggctcgccga taggggtgcg cttcgcgtac tccaacacct 601 gctgccacac cagttcgtca tcgtcggccc gcagctcgac gccggtgtag gtgatcttca 661 cgtccttgtt gacgtggaaa atgaccttgt ttttcagcgc ctcgcgcggg attttcttgt 721 tgcgcgtggt gaacagggca gagcgggccg tgtcgtttgg catcgctcgc atcgtgtccg 781 gccacggcgc aatatcgaac aaggaaagct gcatttcctt gatctgctgc ttcgtgtgtt 841 tcagcaacgc ggcctgcttg gcttcgctga cctgttttgc caggtcctcg ccggcggttt 901 ttcgcttctt ggtcgtcata gttcctcgcg tgtcgatggt catcgacttc gccaaacctg 961 ccgcctcctg ttcgagacga cgcgaacgct ccacggcggc cgatggcgcg ggcagggcag 1021 ggggagccag ttgcacgctg tcgcgctcga tcttggccgt agcttgctgg actatcgagc 1081 cgacggactg gaaggtttcg cggggcgcac gcatgacggt gcggcttgcg atggtttcgg 1141 catcctcggc ggaaaacccc gcgtcgatca gttcttgcct gtatgccttc cggtcaaacg 1201 tccgattcat tcaccctcct tgcgggattg ccccggaatt aattccccgg atcgatccgt 1261 cgatcttgat cccctgcgcc atcagatcct tggcggcaag aaagccatcc agtttacttt 1321 gcagggcttc ccaaccttac cagagggcgc cccagctggc aattccggtt cgcttgctgt 1381 ccataaaacc gcccagtcta gctatcgcca tgtaagccca ctgcaagcta cctgctttct 1441 ctttgcgctt gcgttttccc ttgtccagat agcccagtag ctgacattca tccggggtca 1501 gcaccgtttc tgcggactgg ctttctacgt ggctgccatt tttggggtga ggccgttcgc 1561 ggccgagggg cgcagcccct ggggggatgg gaggcccgcg ttagcgggcc gggagggttc 1621 gagaaggggg ggcacccccc ttcggcgtgc gcggtcacgc gcacagggcg cagccctggt 1681 taaaaacaag gtttataaat attggtttaa aagcaggtta aaagacaggt tagcggtggc 1741 cgaaaaacgg gcggaaaccc ttgcaaatgc tggattttct gcctgtggac agcccctcaa 1801 atgtcaatag gtgcgcccct catctgtcag cactctgccc ctcaagtgtc aaggatcgcg 1861 cccctcatct gtcagtagtc gcgcccctca agtgtcaata ccgcagggca cttatcccca 1921 ggcttgtcca catcatctgt gggaaactcg cgtaaaatca ggcgttttcg ccgatttgcg 1981 aggctggcca gctccacgtc gccggccgaa atcgagcctg cccctcatct gtcaacgccg 2041 cgccgggtga gtcggcccct caagtgtcaa cgtccgcccc tcatctgtca gtgagggcca 2101 agttttccgc gaggtatcca caacgccggc ggccctacat ggctctgctg tagtgagtgg 2161 gttgcgctcc ggcagcggtc ctgatccccc gcagaaaaaa aggatctcaa gaagatcctt 2221 tgatcttttc tacactagtg cttggattct caccttggtc tgacagttac caatgcgagg 2281 gggggatcca ccttcgggag cgcctgaagc ccgttctgga cgccctgggg ccgttgaatc 2341 gggatatgca ggccaaggcc gccgcgatca tcaaggccgt gggcgaaaag ctgctgacgg 2401 aacagcggga agtccagcgc cagaaacagg cccagcgcca gcaggaacgc gggcgcgcac 2461 atttccccga aaagtgccac ctgcggcgtt gtgacaattt accgaacaac tccgcggccg 2521 ggaagccgat ctcggcttga acgaattgtt aggtggcggt acttgggtcg atatcaaagt 2581 gcatcacttc ttcccgtatg cccaactttg tatagagagc cactgcggga tcgtcaccgt 2641 aatctgcttg cacgtagatc acataagcac caagcgcgtt ggcctcatgc ttgaggagat 2701 tgatgagcgc ggtggcaatg ccctgcctcc ggtgctctcc ggagactgcg agatcataga 2761 tatagatctc actacgcggc tgctcaaact tgggcagaac gtaagccgcg agagcgccaa 2821 caaccacttc ttggtcgaag gcagcaagcg cgatgaatgt cttactacgg agcaagttcc 2881 cgaggtaatc ggagtccggc tgatgttggg agtaggtggc tacgtctccg aactcacgac 2941 cgaaaagatc aagagcagcc cgcatggatt tgacttggtc agggccgagc ctacatgtgc 3001 gaatgatgcc catacttgag ccacctaact ttgttttagg gcgactgccc tgctgcgtaa 3061 catcgttgct gctgcgtaac atcgttgctg ctccataaca tcaaacatcg acccacggcg 3121 taacgcgctt gctgcttgga tgcccgaggc atagactgta caaaaaaaca gtcataacaa 3181 gccatgaaaa ccgccactgc gccgttacca ccgctgcgtt cggtcaaggt tctggaccag 3241 ttgcgtgagc gcatacgcta cttgcattac agtttacgaa ccgaacaggc ttatgtcaac 3301 tgggttcgtg ccttcatccg tttccacggt gtgcgtccat gggcaaatat tatacgcaag 3361 gcgacaaggt gctgatgccg ctggcgattc aggttcatca tgccgtttgt gatggcttcc 3421 atgtcggcag aatgcttaat gaattacaac agtttttatg catgcgcagt ccagttacgc 3481 tggagtctga ggctcgtcct gaatgatatc tcatttatca tcatcatctt tataatcacc 3541 gctgccgtca cctcctagct gactcaaatc aatgcgtgtt tcataaagac cagtgatgga 3601 ttgatggata agagtggcat ctaaaacttc ttttgtagac gtatatcgtt tacgatcaat 3661 tgttgtatca aaatatttaa aagcagcggg agctccaaga ttcgtcaacg taaataaatg 3721 aataatattt tctgcttgtt cacgtattgg tttgtctcta tgtttgttat atgcactaag 3781 aactttatct aaattggcat ctgctaaaat aacacgctta gaaaattcac tgatttgctc 3841 aataatctca tctaaataat gcttatgctg ctccacaaac aattgttttt gttcgttatc 3901 ttctggacta cccttcaact tttcataatg actagctaaa tataaaaaat tcacatattt 3961 gcttggcaga gccagctcat ttcctttttg taattctccg gcactagcca gcatccgttt 4021 acgaccgttt tctaactcaa aaagactata tttaggtagt ttaatgatta agtctttttt 4081 aacttcctta tatcctttag cttctaaaaa gtcaatcgga tttttttcaa aggaacttct 4141 ttccataatt gtgatcccta gtaactcttt aacggatttt aacttcttcg atttcccttt 4201 ttccacctta gcaaccacta ggactgaata agctaccgtt ggactatcaa aaccaccata 4261 tttttttgga tcccagtctt ttttacgagc aataagcttg tccgaatttc tttttggtaa 4321 aattgactcc ttggagaatc cgcctgtctg tacttctgtt ttcttgacaa tattgacttg 4381 gggcatggac aatactttgc gcactgtggc aaaatctcgc cctttatccc agacaatttc 4441 tccagtttcc ccattagttt cgattagagg gcgtttgcga atctctccat ttgcaagtgt 4501 aatttctgtt ttgaagaagt tcatgatatt agagtaaaag aaatattttg cggttgcttt 4561 gcctatttct tgctcagact tagcaatcat tttacgaaca tcataaactt tataatcacc 4621 atagacaaac tccgattcaa gttttggata tttcttaatc aaagcagttc caacgacggc 4681 atttagatac gcatcatggg catgatggta attgttaatc tcacgtactt tatagaattg 4741 gaaatctttt cggaagtcag aaactaattt agattttaag gtaatcactt taacctctcg 4801 aataagttta tcattttcat cgtatttagt attcatgcga ctatccaaaa tttgtgccac 4861 atgcttagtg atttggcgag tttcaaccaa ttggcgtttg ataaaaccag ctttatcaag 4921 ttcactcaaa cctccacgtt cagctttcgt taaattatca aacttacgtt gagtgattaa 4981 cttggcgttt agaagttgtc tccaatagtt tttcatcttt ttgactactt cttcacttgg 5041 aacgttatcc gatttaccac gatttttatc agaacgcgtt aagaccttat tgtctattga 5101 atcgtcttta aggaaacttt gtggaacaat gtgatcgaca tcataatcac ttaaacgatt 5161 aatatctaat tcttggtcca catacatgtc tcttccattt tggagataat agagatagag 5221 cttttcattt tgcaattgag tattttcaac aggatgctct ttaagaatct gacttcctaa 5281 ttctttgata ccttcttcga ttcgtttcat acgctctcgc gaatttttct ggcccttttg 5341 agttgtctga ttttcacgtg ccatttcaat aacgatattt tctggcttat gccgccccat 5401 tactttgacc aattcatcaa caacttttac agtctgtaaa ataccttttt taatagcagg 5461 gctaccagct aaatttgcaa tatgttcatg taaactatcg ccttgtccag acacttgtgc 5521 tttttgaatg tcttctttaa atgtcaaact atcatcatgg atcagctgca taaaattgcg 5581 attggcaaaa ccatctgatt tcaaaaaatc taatattgtt ttgccagatt gcttatccct 5641 aataccatta atcaattttc gagacaaacg tccccaacca gtataacggc gacgtttaag 5701 ctgtttcatc accttatcat caaagaggtg agcatatgtt ttaagtcttt cctcaatcat 5761 ctccctatct tcaaataagg tcaatgttaa aacaatatcc tctaagatat cttcattttc 5821 ttcattatcc aaaaaatctt tatctttaat aatttttagc aaatcatggt aggtacctaa 5881 tgaagcatta aatctatctt caactcctga aatttcaaca ctatcaaaac attctatttt 5941 tttgaaataa tcttctttta attgcttaac ggttactttt cgatttgttt tgaagagtaa 6001 atcaacaatg gctttcttct gttcacctga aagaaatgct ggttttcgca ttccttcagt 6061 aacatatttg acctttgtca attcgttata aaccgtaaaa tactcataaa gcaaactatg 6121 ttttggtagt actttttcat ttggaagatt tttatcaaag tttgtcatgc gttcaataaa 6181 tgattgagct gaagcacctt tatcgacaac ttcttcaaaa ttccatgggg taattgtttc 6241 ttcagacttc cgagtcatcc atgcaaaacg actattgcca cgcgccaatg gaccaacata 6301 ataaggaatt cgaaaagtca agattttttc aatcttctca cgattgtctt ttaaaaatgg 6361 ataaaagtct tcttgtcttc tcaaaatagc atgcagctca cccaagtgaa tttgatgggg 6421 aatagagccg ttgtcaaagg tccgttgctt gcgcagcaaa tcttcacgat ttagtttcac 6481 caataattcc tcagtaccat ccattttttc taaaattggt ttgataaatt tataaaattc 6541 ttcttggcta gctcccccat caatataacc tgcatatccg ttttttgatt gatcaaaaaa 6601 gatttcttta tacttttctg gaagttgttg tcgaactaaa gcttttaaaa gagtcaagtc 6661 ttgatgatgt tcatcgtagc gtttaatcat tgaagctgat aggggagcct tagttatttc 6721 agtatttact cttaggatat ctgaaagtaa aatagcatct gataaattct tagctgccaa 6781 aaacaaatca gcatattgat ctccaatttg cgccaataaa ttatctaaat catcatcgta 6841 agtatctttt gaaagctgta atttagcatc ttctgccaaa tcaaaatttg atttaaaatt 6901 aggggtcaaa cccaatgaca aagcaatgag attcccaaat aagccatttt tcttctcacc 6961 ggggagctga gcaatgagat tttctaatcg tcttgattta ctcaatcgtg cagaaagaat 7021 cgctttagca tctactccac ttgcgttaat agggttttct tcaaataatt gattgtaggt 7081 ttgtaccaac tggataaata gtttgtccac atcactatta tcaggattta aatctccctc 7141 aatcaaaaaa tgaccacgaa acttaatcat atgcgctaag gccaaataga ttaagcgcaa 7201 atccgcttta tcagtagaat ctaccaattt ttttcgcaga tgatagatag ttggatattt 7261 ctcatgataa gcaacttcat ctactatatt tccaaaaata ggatgacgtt catgcttctt 7321 gtcttcttcc accaaaaaag actcttcaag tcgatgaaag aaactatcat ctactttcgc 7381 catctcattt gaaaaaatct cctgtagata acaaatacga ttcttccgac gtgtatacct 7441 tctacgagct gtccgtttga gacgagtcgc ttccgctgtc tctccactgt caaataaaag 7501 agcccctata agattttttt tgatactgtg gcggtctgta tttcccagaa ccttgaactt 7561 tttagacgga accttatatt catcagtgat caccgcccat ccgacgctat ttgtgccgat 7621 atctaagcct attgagtatt tcttatccat tgattgtacc tcctggtcag tgcgtcctgc 7681 tgatagctag catagtacct aggactgagc tagccgtaaa ctctagaagc ggccgcgaat 7741 tcaactgcag tctagactcg agtaaggatc cagttcaccg acaaacaaca gataaaacga 7801 aaggcccagt ctttcgactg agcctttcgt tttatttgat gcctggcagt tccctactct 7861 cgcatgggga gaccccacac taccatcggc gctacggcgt ttcacttctg agttcggcat 7921 ggggtcaggt gggaccaccg cgctactgcc gccaggcaaa ttctgtttta tcagaccgct 7981 tctgcgttct gatttaatct gtatcaggct gaaaatcttc tctcatccgc caaaacagcc 8041 aagctggaga ccgtttaaac tcaatgatga tgatgatgat ggtcgacggc gctattcaga 8101 tcctcttctg agatgagttt ttgttcgggc ccaagcttca aaaaaagcac cgactcggtg 8161 ccactttttc aagttgataa cggactagcc ttattttaac ttgctatttc tagctctaaa 8221 acggaagctc cctcgtgcgc tcgtgctcag tatctctatc actgataggg atgtcaatct 8281 ctatcactga tagggactcg agctatgcta ctccgtcaag ccgtcaattg tctgattcgt 8341 taccaactcg aggacaccat cgaatggcgc aaaacctttc gcggtatggc atgatagcgc 8401 ccggaagaga gtcaattcag ggtggtgaat atgtctagat tagataaaag taaagtgatt 8461 aacagcgcat tagagctgct taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc 8521 gcccagaagc taggtgtaga gcagcctaca ttgtattggc atgtaaaaaa taagcgggct 8581 ttgctcgacg ccttagccat tgagatgtta gataggcacc atactcactt ttgcccttta 8641 gaaggggaaa gctggcaaga ttttttacgt aataacgcta aaagttttag atgtgcttta 8701 ctaagtcatc gcgatggagc aaaagtacat ttaggtacac ggcctacaga aaaacagtat 8761 gaaactctcg aaaatcaatt agccttttta tgccaacaag gtttttcact agagaatgca 8821 ttatatgcac tcagcgctgt ggggcatttt actttaggtt gcgtattgga agatcaagag 8881 catcaagtcg ctaaagaaga aagggaaaca cctactactg atagtatgcc gccattatta 8941 cgacaagcta tcgaattatt tgatcaccaa ggtgcagagc cagccttctt attcggcctt 9001 gaattgatca tatgcggatt agaaaaacaa cttaaatgtg aaagtgggtc ttaataccac 9061 cgtcaaaaaa aacggcgctt tttagcgccg tttttatttt tcaaccttaa gcttgaattc 9121 gttgacgaat tctctagata tcgctcaata ctgaccattt aaatcatacc tgacctccat 9181 agcagaaagt caaaaacctc cgaccggagg cttttgactt gagggggatc cggcacattt 9241 ccccgaaaag tgccacctgc atcgatttat tatgacaact tgacggctac atcattcact 9301 ttttcttcac aaccggcacg gaactcgctc gggctggccc cggtgcattt tttaaatacc 9361 cgcgagaagt agagttgatc gtcaaaacca acattgcgac cgacggtggc gataggcatc 9421 cgggtggtgc tcaaaagcag cttcgcctgg ctgatacgtt ggtcctcgcg ccagcttaag 9481 acgctaatcc ctaactgctg gcggaaaaga tgtgacagac gcgacggcga caagcaaaca 9541 tgctgtgcga cgctggcgat atcaaaattg ctgtctgcca ggtgatcgct gatgtactga 9601 caagcctcgc gtacccgatt atccatcggt ggatggagcg actcgttaat cgcttccatg 9661 cgccgcagta acaattgctc aagcagattt atcgccagca gctccgaata gcgcccttcc 9721 ccttgcccgg cgttaatgat ttgcccaaac aggtcgctga aatgcggctg gtgcgcttca 9781 tccgggcgaa agaaccccgt attggcaaat attgacggcc agttaagcca ttcatgccag 9841 taggcgcgcg gacgaaagta aacccactgg tgataccatt cgcgagcctc cggatgacga 9901 ccgtagtgat gaatctctcc tggcgggaac agcaaaatat cacccggtcg gcaaacaaat 9961 tctcgtccct gatttttcac caccccctga ccgcgaatgg tgagattgag aatataacct 10021 ttcattccca gcggtcggtc gataaaaaaa tcgagataac cgttggcctc aatcggcgtt 10081 aaacccgcca ccagatgggc attaaacgag tatcccggca gcaggggatc attttgcgct 10141 tcagccatac ttttcatact cccgccattc agagaagaaa ccaattgtcc atattgcatc 10201 agacattgcc gtcactgcgt cttttactgg ctcttctcgc taaccaaacc ggtaaccccg 10261 cttattaaaa gcattctgta acaaagcggg accaaagcca tgacaaaaac gcgtaacaaa 10321 agtgtctata atcacggcag aaaagtccac attgattatt tgcacggcgt cacactttgc 10381 tatgccatag catttttatc cataagatta gcggatccta cctgacgctt tttatcgcaa 10441 ctctctactg tttctccata cccgtttttt tgggaattcg agctctaagg aggttataaa 10501 aaatggatat taatactgaa actgagatca agcaaaagca ttcactaacc ccctttcctg 10561 ttttcctaat cagcccggca tttcgcgggc gatattttca cagctatttc aggagttcag 10621 ccatgaacgc ttattacatt caggatcgtc ttgaggctca gagctgggcg cgtcactacc 10681 agcagctcgc ccgtgaagag aaagaggcag aactggcaga cgacatggaa aaaggcctgc 10741 cccagcacct gtttgaatcg ctatgcatcg atcatttgca acgccacggg gccagcaaaa 10801 aatccattac ccgtgcgttt gatgacgatg ttgagtttca ggagcgcatg gcagaacaca 10861 tccggtacat ggttgaaacc attgctcacc accaggttga tattgattca gaggtataaa 10921 acgaatgagt actgcactcg caacgctggc tgggaagctg gctgaacgtg tcggcatgga 10981 ttctgtcgac ccacaggaac tgatcaccac tcttcgccag acggcattta aaggtgatgc 11041 cagcgatgcg cagttcatcg cattactgat cgttgccaac cagtacggcc ttaatccgtg 11101 gacgaaagaa atttacgcct ttcctgataa gcagaatggc atcgttccgg tggtgggcgt 11161 tgatggctgg tcccgcatca tcaatgaaaa ccagcagttt gatggcatgg actttgagca 11221 ggacaatgaa tcctgtacat gccggattta ccgcaaggac cgtaatcatc cgatctgcgt 11281 taccgaatgg atggatgaat gccgccgcga accattcaaa actcgcgaag gcagagaaat 11341 cacggggccg tggcagtcgc atcccaaacg gatgttacgt cataaagcca tgattcagtg 11401 tgcccgtctg gccttcggat ttgctggtat ctatgacaag gatgaagccg agcgcattgt 11461 cgaaaatact gcatacactg cagaacgtca gccggaacgc gacatcactc cggttaacga 11521 tgaaaccatg caggagatta acactctgct gatcgccctg gataaaacat gggatgacga 11581 cttattgccg ctctgttccc agatatttcg ccgcgacatt cgtgcatcgt cagaactgac 11641 acaggccgaa gcagtaaaag ctcttggatt cctgaaacag aaagccgcag agcagaaggt 11701 ggcagcatga caccggacat tatcctgcag cgtaccggga tcgatgtgag agctgtcgaa 11761 cagggggatg atgcgtggca caaattacgg ctcggcgtca tcaccgcttc agaagttcac 11821 aacgtgatag caaaaccccg ctccggaaag aagtggcctg acatgaaaat gtcctacttc 11881 cacaccctgc ttgctgaggt ttgcaccggt gtggctccgg aagttaacgc taaagcactg 11941 gcctggggaa aacagtacga gaacgacgcc agaaccctgt ttgaattcac ttccggcgtg 12001 aatgttactg aatccccgat catctatcgc gacgaaagta tgcgtaccgc ctgctctccc 12061 gatggtttat gcagtgacgg caacggcctt gaactgaaat gcccgtttac ctcccgggat 12121 ttcatgaagt tccggctcgg tggtttcgag gccataaagt cagcttacat ggcccaggtg 12181 cagtacagca tgtgggtgac gcgaaaaaat gcctggtact ttgccaacta tgacccgcgt 12241 atgaagcgtg aaggcctgca ttatgtcgtg attgagcggg atgaaaagta catggcgagt 12301 tttgacgaga tcgtgccgga gttcatcgaa aaaatggacg aggcactggc tgaaattggt 12361 tttgtatttg gggagcaatg gcgatgacgc atcctcacga taatatccgg gtaggcgcaa 12421 tcactttcgt ctactccgtt acaaagcgag gctgggtatt tcccggcctt tctgttatcc 12481 gaaatccact gaaagcacag cggctggctg aggagataaa taataaacga ggggctgtat 12541 gcacaaagca tcttctgttg agttaagaac gagtatcgag atggcacata gccttgctca 12601 aattggaatc aggtttgtgc caataccagt agaaacagac gaagaatcca tgggtatgga 12661 cagttttccc t //