LOCUS Exported 13567 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses taCas9 in Mammalian cells for transactivating endogenous and synthetic promoters. The backbone is a lentiviral vector.. ACCESSION . VERSION . KEYWORDS CMVp-dCas9-3xNLS-VP64 (Construct 1) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 13567) AUTHORS Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK TITLE Multiplexed and Programmable Regulation of Gene Networks with an Integrated RNA and CRISPR/Cas Toolkit in Human Cells. JOURNAL Mol Cell. 2014 May 14. pii: S1097-2765(14)00355-4. doi: 10.1016/j.molcel.2014.04.022. PUBMED 24837679 REFERENCE 2 (bases 1 to 13567) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..13567 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(47..66) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 238..617 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 619..817 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 772..792 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 835..1015 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 1062..1187 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1680..1913 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 2098..2142 /codon_start=1 /product="antigenic peptide corresponding to amino acids 655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik et al., 2013)" /label=gp41 peptide /note="recognized by the 2H10 single-chain llama nanobody" /translation="KNEQELLELDKWASL" misc_feature 2440..2557 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2611..3821 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 3658..3677 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" regulatory 3824..3833 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 3830..7933 /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" CDS 7970..7990 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 7997..8017 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 8024..8044 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 8084..8233 /codon_start=1 /product="tetrameric repeat of the minimal activation domain of herpes simplex virus VP16 (Beerli et al., 1998)" /label=VP64 /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF DLDML" misc_feature 8259..8847 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(8312..8332) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(8730..8741) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(8850..8866) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(8851..8867) /label=pBluescriptKS /note="For pBluescript vector" LTR 9372..9552 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" primer_bind complement(9578..9595) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 9584..9808 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 9854..10282 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9941..9960) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 10151..10172 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(10291..10311) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 10296..10625 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 10476..10611 /label=SV40 ori /note="SV40 origin of replication" primer_bind 10538..10557 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 10673..10720 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 10739..11113 /codon_start=1 /gene="Sh ble from Streptoalloteichus hindustanus" /product="antibiotic-binding protein" /label=BleoR /note="confers resistance to bleomycin, phleomycin, and Zeocin(TM)" /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR EFALRDPAGNCVHFVAEEQD" polyA_signal 11243..11364 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(11280..11299) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 11334..11353 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(11413..11429) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(11413..11429) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(11426..11448) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 11437..11453 /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(11461..11491) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 11506..11527 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(11644..11661) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(11815..12403) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(11895..11914) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(12574..13434) /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 13197..13216 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(13435..13539) /gene="bla" /label=AmpR promoter ORIGIN 1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg 61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt 121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc 181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac 241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat 301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg 361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt 421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag 481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc 541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag 601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt 661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc 721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg 781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct 841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt 901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac 961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc 1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc 1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa 1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg 1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata 1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc 1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga 1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc 1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca 1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg 1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga 1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata 1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg 1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg 1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag 1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt 1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt 1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt 2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag 2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata 2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta 2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta 2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa 2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt 2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat 2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa 2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag 2581 agatccagtt tggttaatta agggtgcagc ggcctccgcg ccgggttttg gcgcctcccg 2641 cgggcgcccc cctcctcacg gcgagcgctg ccacgtcaga cgaagggcgc aggagcgttc 2701 ctgatccttc cgcccggacg ctcaggacag cggcccgctg ctcataagac tcggccttag 2761 aaccccagta tcagcagaag gacattttag gacgggactt gggtgactct agggcactgg 2821 ttttctttcc agagagcgga acaggcgagg aaaagtagtc ccttctcggc gattctgcgg 2881 agggatctcc gtggggcggt gaacgccgat gattatataa ggacgcgccg ggtgtggcac 2941 agctagttcc gtcgcagccg ggatttgggt cgcggttctt gtttgtggat cgctgtgatc 3001 gtcacttggt gagttgcggg ctgctgggct ggccggggct ttcgtggccg ccgggccgct 3061 cggtgggacg gaagcgtgtg gagagaccgc caagggctgt agtctgggtc cgcgagcaag 3121 gttgccctga actgggggtt ggggggagcg cacaaaatgg cggctgttcc cgagtcttga 3181 atggaagacg cttgtaaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg 3241 gcggcaagaa cccaaggtct tgaggccttc gctaatgcgg gaaagctctt attcgggtga 3301 gatgggctgg ggcaccatct ggggaccctg acgtgaagtt tgtcactgac tggagaactc 3361 gggtttgtcg tctggttgcg ggggcggcag ttatgcggtg ccgttgggca gtgcacccgt 3421 acctttggga gcgcgcgcct cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg 3481 cagggtgggg ccacctgccg gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg 3541 gttcgggcct agggtaggct ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg 3601 gataagtgag gcgtcagttt ctttggtcgg ttttatgtac ctatcttctt aagtagctga 3661 agctccggtt ttgaactatg cgctcggggt tggcgagtgt gttttgtgaa gttttttagg 3721 caccttttga aatgtaatca tttgggtcaa tatgtaattt tcagtgttag actagtaaat 3781 tgtccgctaa attctggccg tttttggctt ttttgttaga caggccacca tggacaagaa 3841 gtactccatt gggctcgcca tcggcacaaa cagcgtcggc tgggccgtca ttacggacga 3901 gtacaaggtg ccgagcaaaa aattcaaagt tctgggcaat accgatcgcc acagcataaa 3961 gaagaacctc attggcgccc tcctgttcga ctccggggag acggccgaag ccacgcggct 4021 caaaagaaca gcacggcgca gatatacccg cagaaagaat cggatctgct acctgcagga 4081 gatctttagt aatgagatgg ctaaggtgga tgactctttc ttccataggc tggaggagtc 4141 ctttttggtg gaggaggata aaaagcacga gcgccaccca atctttggca atatcgtgga 4201 cgaggtggcg taccatgaaa agtacccaac catatatcat ctgaggaaga agcttgtaga 4261 cagtactgat aaggctgact tgcggttgat ctatctcgcg ctggcgcata tgatcaaatt 4321 tcggggacac ttcctcatcg agggggacct gaacccagac aacagcgatg tcgacaaact 4381 ctttatccaa ctggttcaga cttacaatca gcttttcgaa gagaacccga tcaacgcatc 4441 cggagttgac gccaaagcaa tcctgagcgc taggctgtcc aaatcccggc ggctcgaaaa 4501 cctcatcgca cagctccctg gggagaagaa gaacggcctg tttggtaatc ttatcgccct 4561 gtcactcggg ctgaccccca actttaaatc taacttcgac ctggccgaag atgccaagct 4621 tcaactgagc aaagacacct acgatgatga tctcgacaat ctgctggccc agatcggcga 4681 ccagtacgca gacctttttt tggcggcaaa gaacctgtca gacgccattc tgctgagtga 4741 tattctgcga gtgaacacgg agatcaccaa agctccgctg agcgctagta tgatcaagcg 4801 ctatgatgag caccaccaag acttgacttt gctgaaggcc cttgtcagac agcaactgcc 4861 tgagaagtac aaggaaattt tcttcgatca gtctaaaaat ggctacgccg gatacattga 4921 cggcggagca agccaggagg aattttacaa atttattaag cccatcttgg aaaaaatgga 4981 cggcaccgag gagctgctgg taaagcttaa cagagaagat ctgttgcgca aacagcgcac 5041 tttcgacaat ggaagcatcc cccaccagat tcacctgggc gaactgcacg ctatcctcag 5101 gcggcaagag gatttctacc cctttttgaa agataacagg gaaaagattg agaaaatcct 5161 cacatttcgg ataccctact atgtaggccc cctcgcccgg ggaaattcca gattcgcgtg 5221 gatgactcgc aaatcagaag agaccatcac tccctggaac ttcgaggaag tcgtggataa 5281 gggggcctct gcccagtcct tcatcgaaag gatgactaac tttgataaaa atctgcctaa 5341 cgaaaaggtg cttcctaaac actctctgct gtacgagtac ttcacagttt ataacgagct 5401 caccaaggtc aaatacgtca cagaagggat gagaaagcca gcattcctgt ctggagagca 5461 gaagaaagct atcgtggacc tcctcttcaa gacgaaccgg aaagttaccg tgaaacagct 5521 caaagaagac tatttcaaaa agattgaatg tttcgactct gttgaaatca gcggagtgga 5581 ggatcgcttc aacgcatccc tgggaacgta tcacgatctc ctgaaaatca ttaaagacaa 5641 ggacttcctg gacaatgagg agaacgagga cattcttgag gacattgtcc tcacccttac 5701 gttgtttgaa gatagggaga tgattgaaga acgcttgaaa acttacgctc atctcttcga 5761 cgacaaagtc atgaaacagc tcaagaggcg ccgatataca ggatgggggc ggctgtcaag 5821 aaaactgatc aatgggatcc gagacaagca gagtggaaag acaatcctgg attttcttaa 5881 gtccgatgga tttgccaacc ggaacttcat gcagttgatc catgatgact ctctcacctt 5941 taaggaggac atccagaaag cacaagtttc tggccagggg gacagtcttc acgagcacat 6001 cgctaatctt gcaggtagcc cagctatcaa aaagggaata ctgcagaccg ttaaggtcgt 6061 ggatgaactc gtcaaagtaa tgggaaggca taagcccgag aatatcgtta tcgagatggc 6121 ccgagagaac caaactaccc agaagggaca gaagaacagt agggaaagga tgaagaggat 6181 tgaagagggt ataaaagaac tggggtccca aatccttaag gaacacccag ttgaaaacac 6241 ccagcttcag aatgagaagc tctacctgta ctacctgcag aacggcaggg acatgtacgt 6301 ggatcaggaa ctggacatca atcggctctc cgactacgac gtggatgcca tcgtgcccca 6361 gtcttttctc aaagatgatt ctattgataa taaagtgttg acaagatccg ataaaaatag 6421 agggaagagt gataacgtcc cctcagaaga agttgtcaag aaaatgaaaa attattggcg 6481 gcagctgctg aacgccaaac tgatcacaca acggaagttc gataatctga ctaaggctga 6541 acgaggtggc ctgtctgagt tggataaagc cggcttcatc aaaaggcagc ttgttgagac 6601 acgccagatc accaagcacg tggcccaaat tctcgattca cgcatgaaca ccaagtacga 6661 tgaaaatgac aaactgattc gagaggtgaa agttattact ctgaagtcta agctggtctc 6721 agatttcaga aaggactttc agttttataa ggtgagagag atcaacaatt accaccatgc 6781 gcatgatgcc tacctgaatg cagtggtagg cactgcactt atcaaaaaat atcccaagct 6841 tgaatctgaa tttgtttacg gagactataa agtgtacgat gttaggaaaa tgatcgcaaa 6901 gtctgagcag gaaataggca aggccaccgc taagtacttc ttttacagca atattatgaa 6961 ttttttcaag accgagatta cactggccaa tggagagatt cggaagcgac cacttatcga 7021 aacaaacgga gaaacaggag aaatcgtgtg ggacaagggt agggatttcg cgacagtccg 7081 gaaggtcctg tccatgccgc aggtgaacat cgttaaaaag accgaagtac agaccggagg 7141 cttctccaag gaaagtatcc tcccgaaaag gaacagcgac aagctgatcg cacgcaaaaa 7201 agattgggac cccaagaaat acggcggatt cgattctcct acagtcgctt acagtgtact 7261 ggttgtggcc aaagtggaga aagggaagtc taaaaaactc aaaagcgtca aggaactgct 7321 gggcatcaca atcatggagc gatcaagctt cgaaaaaaac cccatcgact ttctcgaggc 7381 gaaaggatat aaagaggtca aaaaagacct catcattaag cttcccaagt actctctctt 7441 tgagcttgaa aacggccgga aacgaatgct cgctagtgcg ggcgagctgc agaaaggtaa 7501 cgagctggca ctgccctcta aatacgttaa tttcttgtat ctggccagcc actatgaaaa 7561 gctcaaaggg tctcccgaag ataatgagca gaagcagctg ttcgtggaac aacacaaaca 7621 ctaccttgat gagatcatcg agcaaataag cgaattctcc aaaagagtga tcctcgccga 7681 cgctaacctc gataaggtgc tttctgctta caataagcac agggataagc ccatcaggga 7741 gcaggcagaa aacattatcc acttgtttac tctgaccaac ttgggcgcgc ctgcagcctt 7801 caagtacttc gacaccacca tagacagaaa gcggtacacc tctacaaagg aggtcctgga 7861 cgccacactg attcatcagt caattacggg gctctatgaa acaagaatcg acctctctca 7921 gctcggtgga gacagcaggg ctgacgggcc ctcactgggt tcagggtcac ccaagaagaa 7981 acgcaaagtc gaggatccaa agaagaaaag gaaggttgaa gaccccaaga aaaagaggaa 8041 ggtggatggg atcggctcag gcagcaacgg cggtggaggt tcagacgctt tggacgattt 8101 cgatctcgat atgctcggtt ctgacgccct ggatgatttc gatctggata tgctcggcag 8161 cgacgctctc gacgatttcg acctcgacat gctcgggtca gatgccttgg atgattttga 8221 cctggatatg ctctcatgac gatatcaagc ttatcgataa tcaacctctg gattacaaaa 8281 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg 8341 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct 8401 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg 8461 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct 8521 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg 8581 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg 8641 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc 8701 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc 8761 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc 8821 ggatctccct ttgggccgcc tccccgcatc gataccgtcg acctcgagac ctagaaaaac 8881 atggagcaat cacaagtagc aatacagcag ctaccaatgc tgattgtgcc tggctagaag 8941 cacaagagga ggaggaggtg ggttttccag tcacacctca ggtaccttta agaccaatga 9001 cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc 9061 taattcactc ccaacgaaga caagatatcc ttgatctgtg gatctaccac acacaaggct 9121 acttccctga ttggcagaac tacacaccag ggccagggat cagatatcca ctgacctttg 9181 gatggtgcta caagctagta ccagttgagc aagagaaggt agaagaagcc aatgaaggag 9241 agaacacccg cttgttacac cctgtgagcc tgcatgggat ggatgacccg gagagagaag 9301 tattagagtg gaggtttgac agccgcctag catttcatca catggcccga gagctgcatc 9361 cggactgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac 9421 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg 9481 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga 9541 aaatctctag cagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc 9601 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc 9661 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct 9721 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg 9781 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct 9841 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg 9901 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct 9961 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta 10021 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt 10081 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg 10141 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat 10201 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt 10261 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt 10321 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca 10381 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt 10441 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt 10501 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg 10561 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt 10621 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcagcac gtgttgacaa 10681 ttaatcatcg gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat 10741 ggccaagttg accagtgccg ttccggtgct caccgcgcgc gacgtcgccg gagcggtcga 10801 gttctggacc gaccggctcg ggttctcccg ggacttcgtg gaggacgact tcgccggtgt 10861 ggtccgggac gacgtgaccc tgttcatcag cgcggtccag gaccaggtgg tgccggacaa 10921 caccctggcc tgggtgtggg tgcgcggcct ggacgagctg tacgccgagt ggtcggaggt 10981 cgtgtccacg aacttccggg acgcctccgg gccggccatg accgagatcg gcgagcagcc 11041 gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac tgcgtgcact tcgtggccga 11101 ggagcaggac tgacacgtgc tacgagattt cgattccacc gccgccttct atgaaaggtt 11161 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat 11221 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag 11281 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt 11341 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt 11401 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca 11461 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact 11521 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct 11581 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc 11641 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca 11701 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg 11761 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca 11821 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa 11881 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc 11941 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc 12001 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct 12061 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg 12121 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag 12181 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta 12241 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg 12301 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt 12361 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt 12421 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag 12481 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat 12541 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc 12601 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat 12661 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc 12721 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag 12781 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag 12841 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt 12901 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg 12961 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt 13021 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc 13081 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc 13141 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa 13201 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg 13261 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc 13321 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag 13381 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt 13441 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt 13501 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc 13561 acctgac //