LOCUS Exported 15709 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION 2nd Generation Lentiviral vector. Expresses an N-terminal p300-dCas9 fusion protein and mCherry. ACCESSION . VERSION . KEYWORDS pHR_TRE3G-p300-dCas9-P2A-mCherry SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 15709) AUTHORS Li K, Liu Y, Cao H, Zhang Y, Gu Z, Liu X, Yu A, Kaphle P, Dickerson KE, Ni M, Xu J TITLE Interrogation of enhancer function by enhancer-targeting CRISPR epigenetic editing. JOURNAL Nat Commun. 2020 Jan 24;11(1):485. doi: 10.1038/s41467-020-14362-5. PUBMED 31980609 REFERENCE 2 (bases 1 to 15709) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..15709 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(1..17) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2..18) /label=pBluescriptKS /note="For pBluescript vector" LTR 108..341 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" promoter complement(366..384) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(367..384) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(674..693) /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" primer_bind complement(907..926) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 1026..1048 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(1086..1104) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 1172..1276 /gene="bla" /label=AmpR promoter CDS 1277..2137 /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 complement(1495..1514) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 2308..2896 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 2797..2816 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 3050..3067 /label=L4440 /note="L4440 vector, forward primer" primer_bind complement(3137..3157) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3142..3471 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3322..3457 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3384..3403 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" intron 4605..4670 /label=small t intron /note="SV40 (simian virus 40) small t antigen intron" CDS 4800..4820 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" polyA_signal 5245..5379 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5282..5301) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5336..5355 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" LTR 5548..6181 /label=3' LTR /note="3' long terminal repeat (LTR) from HIV-1" misc_feature 6228..6353 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 6850..7082 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 7267..7311 /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 7596..7713 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 7775..8124 /label=TRE3GV promoter /note="3rd-generation Tet-responsive promoter that can be activated by binding of Tet-On(R) 3G, optimized for retroviral and lentiviral vectors" protein_bind 7783..7801 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 7819..7837 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 7855..7873 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 7891..7909 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 7927..7945 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 7963..7981 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 7999..8017 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" primer_bind 8063..8087 /label=LNCX /note="Human CMV promoter, forward primer" CDS 8181..10031 /codon_start=1 /gene="EP300" /product="catalytic histone acetyltransferase core of the human E1A-associated protein p300 (Delvecchio et al., 2013)" /label=p300 core /translation="IFKPEELRQALMPTLEALYRQDPESLPFRQPVDPQLLGIPDYFDI VKSPMDLSTIKRKLDTGQYQEPWQYVDDIWLMFNNAWLYNRKTSRVYKYCSKLSEVFEQ EIDPVMQSLGYCCGRKLEFSPQTLCCYGKQLCTIPRDATYYSYQNRYHFCEKCFNEIQG ESVSLGDDPSQPQTTINKEQFSKRKNDTLDPELFVECTECGRKMHQICVLHHEIIWPAG FVCDGCLKKSARTRKENKFSAKRLPSTRLGTFLENRVNDFLRRQNHPESGEVTVRVVHA SDKTVEVKPGMKARFVDSGEMAESFPYRTKALFAFEEIDGVDLCFFGMHVQEYGSDCPP PNQRRVYISYLDSVHFFRPKCLRTAVYHEILIGYLEYVKKLGYTTGHIWACPPSEGDDY IFHCHPPDQKIPKPKRLQEWYKKMLDKAVSERIVHDYKDIFKQATEDRLTSAKELPYFE GDFWPNVLEESIKELEQEEEERKREENTSNESTDVTKGDSKNAKKKNNKKTSKNKSSLS RGNKKKPGMPNVSNDLSQKLYATMEKHKEVFFVIRLIAGPAANSLPPIVDPDPLIPCDL MDGRDAFLTLARDKHLEFSSLRRAQWSTMCMLVELHTQSQD" regulatory 9795..9804 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 10053..14156 /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 14160..14186 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 14205..14225 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 14232..14252 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 14292..14348 /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" CDS 14349..15059 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(14494..14512) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 14762..14781 /label=mCherry-F /note="mCherry, forward primer" misc_feature 15119..15707 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(15172..15192) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(15590..15601) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" ORIGIN 1 cgataccgtc gacctcgagg gaattaattc gagctcggta cctttaagac caatgactta 61 caaggcagct gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat 121 tcactcccaa cgaagacaag atctgctttt tgcttgtact gggtctctct ggttagacca 181 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag 241 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag 301 atccctcaga cccttttagt cagtgtggaa aatctctagc agcatctaga attaattccg 361 tgtattctat agtgtcacct aaatcgtatg tgtatgatac ataaggttat gtattaattg 421 tagccgcgtt ctaacgacaa tatgtacaag cctaattgtg tagcatctgg cttactgaag 481 cagaccctat catctctctc gtaaactgcc gtcagagtcg gtttggttgg acgaaccttc 541 tgagtttctg gtaacgccgt cccgcacccg gaaatggtca gcgaaccaat cagcagggtc 601 atcgctagcc agatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt 661 gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc 721 gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg 781 ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac 841 ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg aatggtgcac 901 tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc 961 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac 1021 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg 1081 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta 1141 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta 1201 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata 1261 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc 1321 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga 1381 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct 1441 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg 1501 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta 1561 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat 1621 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt 1681 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga 1741 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga 1801 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga 1861 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc 1921 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc 1981 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg 2041 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat 2101 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata 2161 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct 2221 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga 2281 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg 2341 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc 2401 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtctttct 2461 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc 2521 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt 2581 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg 2641 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct 2701 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag 2761 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag 2821 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg 2881 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg 2941 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac 3001 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt 3061 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat 3121 tcattaatgc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca 3181 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc 3241 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata 3301 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg 3361 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag 3421 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagcttgga 3481 cacaagacag gcttgcgaga tatgtttgag aataccactt tatcccgcgt cagggagagg 3541 cagtgcgtaa aaagacgcgg actcatgtga aatactggtt tttagtgcgc cagatctcta 3601 taatctcgcg caacctattt tcccctcgaa cactttttaa gccgtagata aacaggctgg 3661 gacacttcac atgagcgaaa aatacatcgt cacctgggac atgttgcaga tccatgcacg 3721 taaactcgca agccgactga tgccttctga acaatggaaa ggcattattg ccgtaagccg 3781 tggcggtctg taccgggtgc gttactggcg cgtgaactgg gtattcgtca tgtcgatacc 3841 gtttgtattt ccagctacga tcacgacaac cagcgcgagc ttaaagtgct gaaacgcgca 3901 gaaggcgatg gcgaaggctt catcgttatt gatgacctgg tggataccgg tggtactgcg 3961 gttgcgattc gtgaaatgta tccaaaagcg cactttgtca ccatcttcgc aaaaccggct 4021 ggtcgtccgc tggttgatga ctatgttgtt gatatcccgc aagatacctg gattgaacag 4081 ccgtgggata tgggcgtcgt attcgtcccg ccaatctccg gtcgctaatc ttttcaacgc 4141 ctggcactgc cgggcgttgt tctttttaac ttcaggcggg ttacaatagt ttccagtaag 4201 tattctggag gctgcatcca tgacacaggc aaacctgagc gaaaccctgt tcaaaccccg 4261 ctttaaacat cctgaaacct cgacgctagt ccgccgcttt aatcacggcg cacaaccgcc 4321 tgtgcagtcg gcccttgatg gtaaaaccat ccctcactgg tatcgcatga ttaaccgtct 4381 gatgtggatc tggcgcggca ttgacccacg cgaaatcctc gacgtccagg cacgtattgt 4441 gatgagcgat gccgaacgta ccgacgatga tttatacgat acggtgattg gctaccgtgg 4501 cggcaactgg atttatgagt gggccccgga tctttgtgaa ggaaccttac ttctgtggtg 4561 tgacataatt ggacaaacta cctacagaga tttaaagctc taaggtaaat ataaaatttt 4621 taagtgtata atgtgttaaa ctactgattc taattgtttg tgtattttag attccaacct 4681 atggaactga tgaatgggag cagtggtgga atgcctttaa tgaggaaaac ctgttttgct 4741 cagaagaaat gccatctagt gatgatgagg ctactgctga ctctcaacat tctactcctc 4801 caaaaaagaa gagaaaggta gaagacccca aggactttcc ttcagaattg ctaagttttt 4861 tgagtcatgc tgtgtttagt aatagaactc ttgcttgctt tgctatttac accacaaagg 4921 aaaaagctgc actgctatac aagaaaatta tggaaaaata ttctgtaacc tttataagta 4981 ggcataacag ttataatcat aacatactgt tttttcttac tccacacagg catagagtgt 5041 ctgctattaa taactatgct caaaaattgt gtacctttag ctttttaatt tgtaaagggg 5101 ttaataagga atatttgatg tatagtgcct tgactagaga tcataatcag ccataccaca 5161 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat 5221 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa 5281 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt 5341 ttgtccaaac tcatcaatgt atcttatcat gtctggatca actggataac tcaagctaac 5401 caaaatcatc ccaaacttcc caccccatac cctattacca ctgccaatta cctagtggtt 5461 tcatttactc taaacctgtg attcctctga attattttca ttttaaagaa attgtatttg 5521 ttaaatatgt actacaaact tagtagttgg aagggctaat tcactcccaa agaagacaag 5581 atatccttga tctgtggatc taccacacac aaggctactt ccctgattag cagaactaca 5641 caccagggcc aggggtcaga tatccactga cctttggatg gtgctacaag ctagtaccag 5701 ttgagccaga taaggtagaa gaggccaata aaggagagaa caccagcttg ttacaccctg 5761 tgagcctgca tgggatggat gacccggaga gagaagtgtt agagtggagg tttgacagcc 5821 gcctagcatt tcatcacgtg gcccgagagc tgcatccgga gtacttcaag aactgctgat 5881 atcgagcttg ctacaaggga ctttccgctg gggactttcc agggaggcgt ggcctgggcg 5941 ggactgggga gtggcgagcc ctcagatcct gcatataagc agctgctttt tgcctgtact 6001 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca 6061 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg 6121 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc 6181 agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca 6241 ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc 6301 caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta 6361 agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa 6421 aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc 6481 ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc 6541 ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg 6601 tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc 6661 aaaacaaaag taagaccacc gcacagcaag cggccggccg ctgatcttca gacctggagg 6721 aggagatatg agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga 6781 accattagga gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc 6841 agtgggaata ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc 6901 agcgtcaatg acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca 6961 gaacaatttg ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg 7021 catcaagcag ctccaggcaa gaatcctggc tgtgaaagat acctaaagga tcaacagctc 7081 ctggggattt ggggttgctc tggaaaactc atttgcacca ctgctgtgcc ttggaatgct 7141 agttggagta ataaatctct ggaacagatt tggaatcaca cgacctggat ggagtgggac 7201 agagaaatta acaattacac aagcttaata cactccttaa ttgaagaatc gcaaaaccag 7261 caagaaaaga atgaacaaga attattggaa ttagataaat gggcaagttt gtggaattgg 7321 tttaacataa caaattggct gtggtatata aaattattca taatgatagt aggaggcttg 7381 gtaggtttaa gaatagtttt tgctgtactt tctatagtga atagagttag gcagggatat 7441 tcaccattat cgtttcagac ccacctccca accccgaggg gacccgacag gcccgaagga 7501 atagaagaag aaggtggaga gagagacaga gacagatcca ttcgattagt gaacggatct 7561 cgacggtatc gccaaatggc agtattcatc cacaatttta aaagaaaagg ggggattggg 7621 gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca aactaaagaa 7681 ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga cagcagagat 7741 ccagtttatc gatgaggccc tttcgtcttc actcgagttt actccctatc agtgatagag 7801 aacgtatgaa gagtttactc cctatcagtg atagagaacg tatgcagact ttactcccta 7861 tcagtgatag agaacgtata aggagtttac tccctatcag tgatagagaa cgtatgacca 7921 gtttactccc tatcagtgat agagaacgta tctacagttt actccctatc agtgatagag 7981 aacgtatatc cagtttactc cctatcagtg atagagaacg tatgtcgagg taggcgtgta 8041 cggtgggcgc ctataaaagc agagctcgtt tagtgaaccg tcagatcgcc tggagcaatt 8101 ccacaacact tttgtcttat acttacgcgt ctgcaggata tcaagcttgc ggtaccgcgg 8161 gcccgggatc cgccaccatg attttcaaac cagaagaact acgacaggca ctgatgccaa 8221 ctttggaggc actttaccgt caggatccag aatcccttcc ctttcgtcaa cctgtggacc 8281 ctcagctttt aggaatccct gattactttg atattgtgaa gagccccatg gatctttcta 8341 ccattaagag gaagttagac actggacagt atcaggagcc ctggcagtat gtcgatgata 8401 tttggcttat gttcaataat gcctggttat ataaccggaa aacatcacgg gtatacaaat 8461 actgctccaa gctctctgag gtctttgaac aagaaattga cccagtgatg caaagccttg 8521 gatactgttg tggcagaaag ttggagttct ctccacagac actgtgttgc tacggcaaac 8581 agttgtgcac aatacctcgt gatgccactt attacagtta ccagaacagg tatcatttct 8641 gtgagaagtg tttcaatgag atccaagggg agagcgtttc tttgggggat gacccttccc 8701 agcctcaaac tacaataaat aaagaacaat tttccaagag aaaaaatgac acactggatc 8761 ctgaactgtt tgttgaatgt acagagtgcg gaagaaagat gcatcagatc tgtgtccttc 8821 accatgagat catctggcct gctggattcg tctgtgatgg ctgtttaaag aaaagtgcac 8881 gaactaggaa agaaaataag ttttctgcta aaaggttgcc atctaccaga cttggcacct 8941 ttctagagaa tcgtgtgaat gactttctga ggcgacagaa tcaccctgag tcaggagagg 9001 tcactgttag agtagttcat gcttctgaca aaaccgtgga agtaaaacca ggcatgaaag 9061 caaggtttgt ggacagtgga gagatggcag aatcctttcc ataccgaacc aaagccctct 9121 ttgcctttga agaaattgat ggtgttgacc tgtgcttctt tggcatgcat gttcaagagt 9181 atggctctga ctgccctcca cccaaccaga ggagagtata catatcttac ctcgatagtg 9241 ttcatttctt ccgtcctaaa tgcttgagga ctgcagtcta tcatgaaatc ctaattggat 9301 atttagaata tgtcaagaaa ttaggttaca caacagggca tatttgggca tgtccaccaa 9361 gtgagggaga tgattatatc ttccattgcc atcctcctga ccagaagata cccaagccca 9421 agcgactgca ggaatggtac aaaaaaatgc ttgacaaggc tgtatcagag cgtattgtcc 9481 atgactacaa ggatattttt aaacaagcta ctgaagatag attaacaagt gcaaaggaat 9541 tgccttattt cgagggtgat ttctggccca atgttctgga agaaagcatt aaggaactgg 9601 aacaggagga agaagagaga aaacgagagg aaaacaccag caatgaaagc acagatgtga 9661 ccaagggaga cagcaaaaat gctaaaaaga agaataataa gaaaaccagc aaaaataaga 9721 gcagcctgag taggggcaac aagaagaaac ccgggatgcc caatgtatct aacgacctct 9781 cacagaaact atatgccacc atggagaagc ataaagaggt cttctttgtg atccgcctca 9841 ttgctggccc tgctgccaac tccctgcctc ccattgttga tcctgatcct ctcatcccct 9901 gcgatctgat ggatggtcgg gatgcgtttc tcacgctggc aagggacaag cacctggagt 9961 tctcttcact ccgaagagcc cagtggtcca ccatgtgcat gctggtggag ctgcacacgc 10021 agagccagga cacgcgttcc ggcggagggt cgatggacaa gaagtacagc atcggcctgg 10081 ccatcggcac caactctgtg ggctgggccg tgatcaccga cgagtacaag gtgcccagca 10141 agaaattcaa ggtgctgggc aacaccgacc ggcacagcat caagaagaac ctgatcggcg 10201 ccctgctgtt cgacagcgga gaaacagccg aggccacccg gctgaagaga accgccagaa 10261 gaagatacac cagacggaag aaccggatct gctatctgca agagatcttc agcaacgaga 10321 tggccaaggt ggacgacagc ttcttccaca gactggaaga gtccttcctg gtggaagagg 10381 ataagaagca cgagcggcac cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg 10441 agaagtaccc caccatctac cacctgagaa agaaactggt ggacagcacc gacaaggccg 10501 acctgcggct gatctatctg gccctggccc acatgatcaa gttccggggc cacttcctga 10561 tcgagggcga cctgaacccc gacaacagcg acgtggacaa gctgttcatc cagctggtgc 10621 agacctacaa ccagctgttc gaggaaaacc ccatcaacgc cagcggcgtg gacgccaagg 10681 ccatcctgtc tgccagactg agcaagagca gacggctgga aaatctgatc gcccagctgc 10741 ccggcgagaa gaagaatggc ctgttcggca acctgattgc cctgagcctg ggcctgaccc 10801 ccaacttcaa gagcaacttc gacctggccg aggatgccaa actgcagctg agcaaggaca 10861 cctacgacga cgacctggac aacctgctgg cccagatcgg cgaccagtac gccgacctgt 10921 ttctggccgc caagaacctg tccgacgcca tcctgctgag cgacatcctg agagtgaaca 10981 ccgagatcac caaggccccc ctgagcgcct ctatgatcaa gagatacgac gagcaccacc 11041 aggacctgac cctgctgaaa gctctcgtgc ggcagcagct gcctgagaag tacaaagaga 11101 ttttcttcga ccagagcaag aacggctacg ccggctacat cgatggcgga gccagccagg 11161 aagagttcta caagttcatc aagcccatcc tggaaaagat ggacggcacc gaggaactgc 11221 tcgtgaagct gaacagagag gacctgctgc ggaagcagcg gaccttcgac aacggcagca 11281 tcccccacca gatccacctg ggagagctgc acgccattct gcggcggcag gaagattttt 11341 acccattcct gaaggacaac cgggaaaaga tcgagaagat cctgaccttc cgcatcccct 11401 actacgtggg ccctctggcc aggggaaaca gcagattcgc ctggatgacc agaaagagcg 11461 aggaaaccat caccccctgg aacttcgagg aagtggtgga caagggcgcc agcgcccaga 11521 gcttcatcga gcggatgacc aacttcgata agaacctgcc caacgagaag gtgctgccca 11581 agcacagcct gctgtacgag tacttcaccg tgtacaacga gctgaccaaa gtgaaatacg 11641 tgaccgaggg aatgagaaag cccgccttcc tgagcggcga gcagaaaaaa gccatcgtgg 11701 acctgctgtt caagaccaac cggaaagtga ccgtgaagca gctgaaagag gactacttca 11761 agaaaatcga gtgcttcgac tccgtggaaa tctccggcgt ggaagatcgg ttcaacgcct 11821 ccctgggcac ataccacgat ctgctgaaaa ttatcaagga caaggacttc ctggacaatg 11881 aggaaaacga ggacattctg gaagatatcg tgctgaccct gacactgttt gaggacagag 11941 agatgatcga ggaacggctg aaaacctatg cccacctgtt cgacgacaaa gtgatgaagc 12001 agctgaagcg gcggagatac accggctggg gcaggctgag ccggaagctg atcaacggca 12061 tccgggacaa gcagtccggc aagacaatcc tggatttcct gaagtccgac ggcttcgcca 12121 acagaaactt catgcagctg atccacgacg acagcctgac ctttaaagag gacatccaga 12181 aagcccaggt gtccggccag ggcgatagcc tgcacgagca cattgccaat ctggccggca 12241 gccccgccat taagaagggc atcctgcaga cagtgaaggt ggtggacgag ctcgtgaaag 12301 tgatgggccg gcacaagccc gagaacatcg tgatcgaaat ggccagagag aaccagacca 12361 cccagaaggg acagaagaac agccgcgaga gaatgaagcg gatcgaagag ggcatcaaag 12421 agctgggcag ccagatcctg aaagaacacc ccgtggaaaa cacccagctg cagaacgaga 12481 agctgtacct gtactacctg cagaatgggc gggatatgta cgtggaccag gaactggaca 12541 tcaaccggct gtccgactac gatgtggacg ctatcgtgcc tcagagcttt ctgaaggacg 12601 actccatcga taacaaagtg ctgactcgga gcgacaagaa ccggggcaag agcgacaacg 12661 tgccctccga agaggtcgtg aagaagatga agaactactg gcgccagctg ctgaatgcca 12721 agctgattac ccagaggaag ttcgacaatc tgaccaaggc cgagagaggc ggcctgagcg 12781 aactggataa ggccggcttc atcaagagac agctggtgga aacccggcag atcacaaagc 12841 acgtggcaca gatcctggac tcccggatga acactaagta cgacgagaac gacaaactga 12901 tccgggaagt gaaagtgatc accctgaagt ccaagctggt gtccgatttc cggaaggatt 12961 tccagtttta caaagtgcgc gagatcaaca actaccacca cgcccacgac gcctacctga 13021 acgccgtcgt gggaaccgcc ctgatcaaaa agtaccctaa gctggaaagc gagttcgtgt 13081 acggcgacta caaggtgtac gacgtgcgga agatgatcgc caagagcgag caggaaatcg 13141 gcaaggctac cgccaagtac ttcttctaca gcaacatcat gaactttttc aagaccgaga 13201 ttaccctggc caacggcgag atccggaagc ggcctctgat cgagacaaac ggcgaaacag 13261 gcgagatcgt gtgggataag ggccgggact ttgccaccgt gcggaaagtg ctgtctatgc 13321 cccaagtgaa tatcgtgaaa aagaccgagg tgcagacagg cggcttcagc aaagagtcta 13381 tcctgcccaa gaggaacagc gacaagctga tcgccagaaa gaaggactgg gaccctaaga 13441 agtacggcgg cttcgacagc cccaccgtgg cctattctgt gctggtggtg gccaaagtgg 13501 aaaagggcaa gtccaagaaa ctgaagagtg tgaaagagct gctggggatc accatcatgg 13561 aaagaagcag cttcgagaag aatcccatcg actttctgga agccaagggc tacaaagaag 13621 tgaaaaagga cctgatcatc aagctgccta agtactccct gttcgagctg gaaaacggcc 13681 ggaagagaat gctggcctct gccggcgaac tgcagaaggg aaacgaactg gccctgccct 13741 ccaaatatgt gaacttcctg tacctggcca gccactatga gaagctgaag ggctcccccg 13801 aggataatga gcagaaacag ctgtttgtgg aacagcacaa acactacctg gacgagatca 13861 tcgagcagat cagcgagttc tccaagagag tgatcctggc cgacgctaat ctggacaagg 13921 tgctgagcgc ctacaacaag cacagagaca agcctatcag agagcaggcc gagaatatca 13981 tccacctgtt taccctgacc aatctgggag cccctgccgc cttcaagtac tttgacacca 14041 ccatcgaccg gaagaggtac accagcacca aagaggtgct ggacgccacc ctgatccacc 14101 agagcatcac cggcctgtac gagacacgga tcgacctgtc tcagctggga ggcgacgcct 14161 atccctatga cgtgcccgat tatgccagcc tgggcagcgg ctcccccaag aaaaaacgca 14221 aggtggaaga tcctaagaaa aagcggaaag tggacggcat tggtagtggg agcaacggca 14281 gcagcggatc cgctactaac ttcagcctgc tgaagcaggc tggagacgtg gaggagaacc 14341 ctggacctat ggtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc 14401 gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg 14461 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc 14521 ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg 14581 tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt 14641 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc 14701 tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg 14761 gccccgtaat gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg 14821 aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact 14881 acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct 14941 acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac 15001 agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtaag 15061 cggccgcgac tctagagtcg acctgcaggc atgcaagctt gatatcaagc ttatcgataa 15121 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc 15181 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat 15241 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg 15301 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg 15361 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat 15421 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt 15481 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc 15541 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa 15601 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg 15661 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcat //