LOCUS Exported 10763 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION A CRISPR activation device with the necessary genes (dcas9 controlled by Ptet, pspF-Delta-HTH::MCP controlled by promoter J23106), and the reporter part (PpspA-2G6 with sfgfp::ASV).. ACCESSION . VERSION . KEYWORDS pLY59 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 10763) AUTHORS TITLE Engineered CRISPRa enables programmable eukaryote-like gene activation in bacteria JOURNAL Nature Communications 10, August 2019 REFERENCE 2 (bases 1 to 10763) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..10763 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 2..59 /label=his operon terminator /note="This putative transcriptin terminator from the E. coli his operon has a 2-bp deletion introduced during synthesis. Its efficiency has not been determined." primer_bind complement(83..100) /label=L4440 /note="L4440 vector, forward primer" CDS complement(362..1312) /codon_start=1 /gene="rep101" /product="RepA protein needed for replication with the pSC101 origin" /label=Rep101 /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI" rep_origin 1360..1582 /label=pSC101 ori /note="low-copy replication origin that requires the Rep101 protein" terminator 2074..2168 /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS complement(2198..3058) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 2821..2840 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(3059..3163) /gene="bla" /label=AmpR promoter primer_bind 3228..3246 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" terminator 3250..3293 /label=bacterial terminator /note="putative bacterial transcription terminator" misc_feature 3296..3317 /label=BioBrick prefix /note="BioBrick prefix for parts that do not start with ""ATG""" CDS complement(3381..7487) /codon_start=1 /product="catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system" /label=dCas9 /note="RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain" /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL DATLIHQSITGLYETRIDLSQLGGD" RBS 7494..7505 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" promoter 7523..7578 /gene="tetR" /label=tetR/tetA promoters /note="overlapping promoters for bacterial tetR and tetA" protein_bind 7524..7542 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" CDS 7597..8220 /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(7670..7689) /label=Tet-R /note="Tetracycline resistance gene, reverse primer" CDS 8235..8246 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" misc_RNA 8452..8501 /label=sTRSV HHRz /note="hammerhead ribozyme from the tobacco ringspot virus satellite RNA (Khvorova et al., 2003)" RBS 8534..8545 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" CDS 8552..9265 /codon_start=1 /product="GFP variant that folds robustly even when fused to poorly folded proteins (Pedelacq et al., 2006)" /label=superfolder GFP /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" CDS 10701..10712 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" misc_feature 10744..1 /label=BioBrick suffix /note="universal suffix for all parts" ORIGIN 1 gtccggcaaa aaagggcaag gtgtcaccac cctgcccttt ttctttaaaa ccgaaaagat 61 tacttcgcgt tatgcaggct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc 121 ggcgagcggt atcagctcac tcaaaggcgg taatctcgag gttacattgt cgatctgttc 181 atggtgaaca gctttgaatg caccaaaaac tcgtaaaagc tctgatgtat ctatcttttt 241 tacaccgttt tcatctgtgc atatggacag ttttcccttt gatatgtaac ggtgaacagt 301 tgttctactt ttgtttgtta gtcttgatgc ttcactgata gatacaagag ccataagaac 361 ctcagatcct tccgtattta gccagtatgt tctctagtgt ggttcgttgt ttttgcgtga 421 gccatgagaa cgaaccattg agatcatact tactttgcat gtcactcaaa aattttgcct 481 caaaactggt gagctgaatt tttgcagtta aagcatcgtg tagtgttttt cttagtccgt 541 tatgtaggta ggaatctgat gtaatggttg ttggtatttt gtcaccattc atttttatct 601 ggttgttctc aagttcggtt acgagatcca tttgtctatc tagttcaact tggaaaatca 661 acgtatcagt cgggcggcct cgcttatcaa ccaccaattt catattgctg taagtgttta 721 aatctttact tattggtttc aaaacccatt ggttaagcct tttaaactca tggtagttat 781 tttcaagcat taacatgaac ttaaattcat caaggctaat ctctatattt gccttgtgag 841 ttttcttttg tgttagttct tttaataacc actcataaat cctcatagag tatttgtttt 901 caaaagactt aacatgttcc agattatatt ttatgaattt ttttaactgg aaaagataag 961 gcaatatctc ttcactaaaa actaattcta atttttcgct tgagaacttg gcatagtttg 1021 tccactggaa aatctcaaag cctttaacca aaggattcct gatttccaca gttctcgtca 1081 tcagctctct ggttgcttta gctaatacac cataagcatt ttccctactg atgttcatca 1141 tctgagcgta ttggttataa gtgaacgata ccgtccgttc tttccttgta gggttttcaa 1201 tcgtggggtt gagtagtgcc acacagcata aaattagctt ggtttcatgc tccgttaagt 1261 catagcgact aatcgctagt tcatttgctt tgaaaacaac taattcagac atacatctca 1321 attggtctag gtgattttaa tcactatacc aattgagatg ggctagtcaa tgataattac 1381 atgtcctttt cctttgagtt gtgggtatct gtaaattctg ctagaccttt gctggaaaac 1441 ttgtaaattc tgctagaccc tctgtaaatt ccgctagacc tttgtgtgtt ttttttgttt 1501 atattcaagt ggttataatt tatagaataa agaaagaata aaaaaagata aaaagaatag 1561 atcccagccc tgtgtataac tcactacttt agtcagttcc gcagtattac aaaaggatgt 1621 cgcaaacgct gtttgctcct ctacaaaaca gaccttaaaa ccctaaaggc ttaagtagca 1681 ccctcgcaag ctcgggcaaa tcgctgaata ttccttttgt ctccgaccat caggcacctg 1741 agtcgctgtc tttttcgtga cattcagttc gctgcgctca cggctctggc agtgaatggg 1801 ggtaaatggc actacaggcg ccttttatgg attcatgcaa ggaaactacc cataatacaa 1861 gaaaagcccg tcacgggctt ctcagggcgt tttatggcgg gtctgctatg tggtgctatc 1921 tgactttttg ctgttcagca gttcctgccc tctgattttc cagtctgacc acttcggatt 1981 atcccgtgac aggtcattca gactggctaa tgcacccagt aaggcagcgg tatcatcaac 2041 aggcttaccc gtcttactgt ccctagtgct tggattctca ccaataaaaa acgcccggcg 2101 gcaaccgagc gttctgaaca aatccagatg gagttctgag gtcattactg gatctatcaa 2161 caggagtcca agcgagctcg taaacttggt ctgacagtta ccaatgctta atcagtgagg 2221 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt 2281 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag 2341 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc 2401 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag 2461 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca 2521 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa 2581 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga 2641 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata 2701 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca 2761 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg 2821 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg 2881 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg 2941 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag 3001 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac 3061 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca 3121 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt cccatggtgc 3181 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca 3241 cgaggcagaa tttcagataa aaaaaatcct tagctttcgc taaggatgat ttctggaatt 3301 cgcggccgct tctagagggg agaccagaaa caaaaaaagg ggagcggttt cccgctcccc 3361 ttcaataatt ggctctagta ttagtcacct cctagctgac tcaaatcaat gcgtgtttca 3421 taaagaccag tgatggattg atggataaga gtggcatcta aaacttcttt tgtagacgta 3481 tatcgtttac gatcaattgt tgtatcaaaa tatttaaaag cagcgggagc tccaagattc 3541 gtcaacgtaa ataaatgaat aatattttct gcttgttcac gtattggttt gtctctatgt 3601 ttgttatatg cactaagaac tttatctaaa ttggcatctg ctaaaataac acgcttagaa 3661 aattcactga tttgctcaat aatctcatct aaataatgct tatgctgctc cacaaacaat 3721 tgtttttgtt cgttatcttc tggactaccc ttcaactttt cataatgact agctaaatat 3781 aaaaaattca catatttgct tggcagagcc agctcatttc ctttttgtaa ttctccggca 3841 ctagccagca tccgtttacg accgttttct aactcaaaaa gactatattt aggtagttta 3901 atgattaagt cttttttaac ttccttatat cctttagctt ctaaaaagtc aatcggattt 3961 ttttcaaagg aacttctttc cataattgtg atccctagta actctttaac ggattttaac 4021 ttcttcgatt tccctttttc caccttagca accactagga ctgaataagc taccgttgga 4081 ctatcaaaac caccatattt ttttggatcc cagtcttttt tacgagcaat aagcttgtcc 4141 gaatttcttt ttggtaaaat tgactccttg gagaatccgc ctgtctgtac ttctgttttc 4201 ttgacaatat tgacttgggg catggacaat actttgcgca ctgtggcaaa atctcgccct 4261 ttatcccaga caatttctcc agtttcccca ttagtttcga ttagagggcg tttgcgaatc 4321 tctccatttg caagtgtaat ttctgttttg aagaagttca tgatattaga gtaaaagaaa 4381 tattttgcgg ttgctttgcc tatttcttgc tcagacttag caatcatttt acgaacatca 4441 taaactttat aatcaccata gacaaactcc gattcaagtt ttggatattt cttaatcaaa 4501 gcagttccaa cgacggcatt tagatacgca tcatgggcat gatggtaatt gttaatctca 4561 cgtactttat agaattggaa atcttttcgg aagtcagaaa ctaatttaga ttttaaggta 4621 atcactttaa cctctcgaat aagtttatca ttttcatcgt atttagtatt catgcgacta 4681 tccaaaattt gtgccacatg cttagtgatt tggcgagttt caaccaattg gcgtttgata 4741 aaaccagctt tatcaagttc actcaaacct ccacgttcag ctttcgttaa attatcaaac 4801 ttacgttgag tgattaactt ggcgtttaga agttgtctcc aatagttttt catctttttg 4861 actacttctt cacttggaac gttatccgat ttaccacgat ttttatcaga acgcgttaag 4921 accttattgt ctattgaatc gtctttaagg aaactttgtg gaacaatggc atcgacatca 4981 taatcactta aacgattaat atctaattct tggtccacat acatgtctct tccattttgg 5041 agataataga gatagagctt ttcattttgc aattgagtat tttcaacagg atgctcttta 5101 agaatctgac ttcctaattc tttgatacct tcttcgattc gtttcatacg ctctcgcgaa 5161 tttttctggc ccttttgagt tgtctgattt tcacgtgcca tttcaataac gatattttct 5221 ggcttatgcc gccccattac tttgaccaat tcatcaacaa cttttacagt ctgtaaaata 5281 ccttttttaa tagcagggct accagctaaa tttgcaatat gttcatgtaa actatcgcct 5341 tgtccagaca cttgtgcttt ttgaatgtct tctttaaatg tcaaactatc atcatggatc 5401 agctgcataa aattgcgatt ggcaaaacca tctgatttca aaaaatctaa tattgttttg 5461 ccagattgct tatccctaat accattaatc aattttcgag acaaacgtcc ccaaccagta 5521 taacggcgac gtttaagctg tttcatcacc ttatcatcaa agaggtgagc atatgtttta 5581 agtctttcct caatcatctc cctatcttca aataaggtca atgttaaaac aatatcctct 5641 aagatatctt cattttcttc attatccaaa aaatctttat ctttaataat ttttagcaaa 5701 tcatggtagg tacctaatga agcattaaat ctatcttcaa ctcctgaaat ttcaacacta 5761 tcaaaacatt ctattttttt gaaataatct tcttttaatt gcttaacggt tacttttcga 5821 tttgttttga agagtaaatc aacaatggct ttcttctgtt cacctgaaag aaatgctggt 5881 tttcgcattc cttcagtaac atatttgacc tttgtcaatt cgttataaac cgtaaaatac 5941 tcataaagca aactatgttt tggtagtact ttttcatttg gaagattttt atcaaagttt 6001 gtcatgcgtt caataaatga ttgagctgaa gcacctttat cgacaacttc ttcaaaattc 6061 catggggtaa ttgtttcttc agacttccga gtcatccatg caaaacgact attgccacgc 6121 gccaatggac caacataata aggaatacga aaagtcaaga ttttttcaat cttctcacga 6181 ttgtctttta aaaatggata aaagtcttct tgtcttctca aaatagcatg cagctcaccc 6241 aagtgaattt gatggggaat agagccgttg tcaaaggtcc gttgcttgcg cagcaaatct 6301 tcacgattta gtttcaccaa taattcctca gtaccatcca ttttttctaa aattggtttg 6361 ataaatttat aaaattcttc ttggctagct cccccatcaa tataacctgc atatccgttt 6421 tttgattgat caaaaaagat ttctttatac ttttctggaa gttgttgtcg aactaaagct 6481 tttaaaagag tcaagtcttg atgatgttca tcgtagcgtt taatcattga agctgatagg 6541 ggagccttag ttatttcagt atttactctt aggatatctg aaagtaaaat agcatctgat 6601 aaattcttag ctgccaaaaa caaatcagca tattgatctc caatttgcgc caataaatta 6661 tctaaatcat catcgtaagt atcttttgaa agctgtaatt tagcatcttc tgccaaatca 6721 aaatttgatt taaaattagg ggtcaaaccc aatgacaaag caatgagatt cccaaataag 6781 ccatttttct tctcaccggg gagctgagca atgagatttt ctaatcgtct tgatttactc 6841 aatcgtgcag aaagaatcgc tttagcatct actccacttg cgttaatagg gttttcttca 6901 aataattgat tgtaggtttg taccaactgg ataaatagtt tgtccacatc actattatca 6961 ggatttaaat ctccctcaat caaaaaatga ccacgaaact taatcatatg cgctaaggcc 7021 aaatagatta agcgcaaatc cgctttatca gtagaatcta ccaatttttt tcgcagatga 7081 tagatagttg gatatttctc atgataagca acttcatcta ctatatttcc aaaaatagga 7141 tgacgttcat gcttcttgtc ttcttccacc aaaaaagact cttcaagtcg atgaaagaaa 7201 ctatcatcta ctttcgccat ctcatttgaa aaaatctcct gtagataaca aatacgattc 7261 ttccgacgtg tataccttct acgagctgtc cgtttgaggc gagtcgcttc cgctgtctct 7321 ccactgtcaa ataaaagagc ccctataaga ttttttttga tactgtggcg gtctgtattt 7381 cccagaacct tgaacttttt agacggaacc ttatattcat cagtgatcac cgcccatccg 7441 acgctatttg tgccgatagc taagcctatt gagtatttct tatccataga tcctttctcc 7501 tctttagatc ttttcaattc ttttctctat cactgatagg gagtggtaaa ataactctat 7561 caacgataga gtgtcaacaa aaattaggaa ttaatgatgt caagattaga taaaagtaaa 7621 gtgattaaca gcgcattaga gctgcttaat gaggtcggaa tcgaaggttt aacaacccgt 7681 aaactcgccc agaagctagg tgtagagcag cctacattgt attggcatgt aaaaaataag 7741 cgggctttgc tcgacgcctt agccattgag atgttagata ggcaccatac tcacttttgc 7801 cctttagaag gggaaagctg gcaagatttt ttacgtaata acgctaaaag ttttagatgt 7861 gctttactaa gtcatcgcga tggagcaaaa gtacatttag gtacacggcc tacagaaaaa 7921 cagtatgaaa ctctcgaaaa tcaattagcc tttttatgcc aacaaggttt ttcactagag 7981 aatgcattat atgcactcag cgctgtgggg cattttactt taggttgcgt attggaagat 8041 caagagcatc aagtcgctaa agaagaaagg gaaacaccta ctactgatag tatgccgcca 8101 ttattacgac aagctatcga attatttgat caccaaggtg cagagccagc cttcttattc 8161 ggccttgaat tgatcatatg cggattagaa aaacaactta aatgtgaaag tgggtcttaa 8221 tactagagcc aattattgaa ggccgctaac gcggcctttt tttgtttctg gtctccctac 8281 tagaaaagct gttgtggcaa aaaacggaaa gctgttgtgg caaaaaacgg catcctggca 8341 tgttgctgtt gattcttcaa tcagatcttt ataaatcaaa aagataaaaa attggcacgc 8401 aaattgtatt aacagttcag caggacaatc ctgaacgcaa atactagaga gctgtcaccg 8461 gatgtgcttt ccggtctgat gagtccgtga ggacgaaaca gcctctacaa ataattttgt 8521 ttaactagag attaaagagg agaaatacca tatgcgtaaa ggcgaagagc tgttcactgg 8581 tgtcgtccct attctggtgg aactggatgg tgatgtcaac ggtcataagt tttccgtgcg 8641 tggcgagggt gaaggtgacg caactaatgg taaactgacg ctgaagttca tctgtactac 8701 tggtaaactg ccggtacctt ggccgactct ggtaacgacg ctgacttatg gtgttcagtg 8761 ctttgctcgt tatccggacc atatgaagca gcatgacttc ttcaagtccg ccatgccgga 8821 aggctatgtg caggaacgca cgatttcctt taaggatgac ggcacgtaca aaacgcgtgc 8881 ggaagtgaaa tttgaaggcg ataccctggt aaaccgcatt gagctgaaag gcattgactt 8941 taaagaagac ggcaatatcc tgggccataa gctggaatac aattttaaca gccacaatgt 9001 ttacatcacc gccgataaac aaaaaaatgg cattaaagcg aattttaaaa ttcgccacaa 9061 cgtggaggat ggcagcgtgc agctggctga tcactaccag caaaacactc caatcggtga 9121 tggtcctgtt ctgctgccag acaatcacta tctgagcacg caaagcgttc tgtctaaaga 9181 tccgaacgag aaacgcgatc atatggttct gctggagttc gtaaccgcag cgggcatcac 9241 gcatggtatg gatgaactgt acaaaaggcc tgctgcaaac gacgaaaact acgctgcatc 9301 agtttaataa tactagagcc aattattgaa ggcctcccta acggggggcc tttttttgtt 9361 tctggtctcc ctactagagt ttacggctag ctcagtccta ggtatagtgc tagctactag 9421 agtcacacag gaaagtacta gatggcagaa tacaaagata atttacttgg tgaggcgaac 9481 agctttctcg aagtgctgga acaggtttcg catctcgcac cgctggacaa accggtgctc 9541 atcatcggcg aacgcggcac cggtaaagag ctgattgcca gccgcctgca ttatctctcc 9601 tcccgttggc aagggccgtt tatttccctt aactgcgcgg cgttaaatga aaatctgctg 9661 gattccgaac tgtttggtca cgaagcgggg gcgtttaccg gtgcgcaaaa acgtcatcca 9721 gggagatttg aacgtgccga cggcggtacg ctatttcttg atgaactcgc tacggcaccg 9781 atgatggtgc aggagaaatt attgcgcgtg attgagtacg gtgaactgga gcgcgttggc 9841 ggcagccaac cattgcaggt gaatgtgcgg ttggtatgcg cgacgaatgc cgatctcccg 9901 gcgatggtca atgaaggcac ttttcgcgct gacctgctcg accgactggc ttttgatgtt 9961 gtacaactgc caccactgcg cgagcgcgaa agcgacataa tgttgatggc agaatacttt 10021 gccatccaga tgtgtcggga aatcaagctg cctctgttcc cggggtttac ggagcgcgcc 10081 agagaaacat tgctgaatta tcgttggccg ggaaatattc gtgaattgaa aaacgtggtg 10141 gaacgttcag tgtatcgcca cggcaccagc gattatccgc ttgatgacat cattattgat 10201 ccctttaaac ggcgtccgcc tgaagacgct atcgccgttt cagaaaccac ctcgcttcca 10261 acactgccgc tggatttacg tgagtttcag atgcagcagg aaaaagagtt gctgcaactc 10321 agtttgcaaa tggcttctaa ctttactcag ttcgttctcg tcgacaatgg cggaactggc 10381 gacgtgactg tcgccccaag caacttcgct aacgggatcg ctgaatggat cagctctaac 10441 tcgcgttcac aggcttacaa agtaacctgt agcgttcgtc agagctctgc gcagaatcgc 10501 aaatacacca tcaaagtcga ggtgcctaaa ggcgcctggc gttcgtactt aaatatggaa 10561 ctaaccattc caattttcgc cacgaactcc gactgcgagc ttattgttaa ggcaatgcaa 10621 ggtctcctaa aagatggaaa cccgattccc tcagcaatcg cagcaaactc cggcatctac 10681 taataatact agagccaatt attgaaggcc gctaacgcgg cctttttttg tttctggtct 10741 ccctactagt agcggccgct gca //