LOCUS Exported 14247 bp ds-DNA circular SYN 23-SEP-2022 DEFINITION 2nd Generation Lentiviral vector. Expresses an N-terminal KRAB-dCas9 fusion protein and mCherry. ACCESSION . VERSION . KEYWORDS pHR-SFFV-KRAB-dCas9-P2A-mCherry SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 14247) AUTHORS Gilbert LA, Horlbeck MA, Adamson B, Villalta JE, Chen Y, Whitehead EH, Guimaraes C, Panning B, Ploegh HL, Bassik MC, Qi LS, Kampmann M, Weissman JS TITLE Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation. JOURNAL Cell. 2014 Oct 23;159(3):647-61. doi: 10.1016/j.cell.2014.09.029. Epub 2014 Oct 9. PUBMED 25307932 REFERENCE 2 (bases 1 to 14247) AUTHORS . TITLE Direct Submission JOURNAL Exported Sep 23, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..14247 /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 5547..6180 /label=3' LTR /note="3' long terminal repeat (LTR) from HIV-1" misc_feature 6227..6352 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 6849..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 7866..8273 /label=SFFV promoter /note="spleen focus-forming virus long terminal repeat (LTR) promoter" regulatory 8348..8357 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 8384..8569 /codon_start=1 /product="Kruppel-associated box (KRAB) transcriptional repression domain from the human zinc finger protein ZNF10 (Margolin et al., 1994)" /label=KRAB /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY QLTKPDVILRLEKGEEP" CDS 8591..12694 /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 12698..12724 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 12743..12763 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 12770..12790 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 12830..12886 /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 12887..13597 /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(13032..13050) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 13300..13319 /label=mCherry-F /note="mCherry, forward primer" misc_feature 13657..14245 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(13710..13730) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(14128..14139) /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 ctgttcttct 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 cctgtggttt 5461 catttactct aaacctgtga ttcctctgaa ttattttcat tttaaagaaa ttgtatttgt 5521 taaatatgta ctacaaactt agtagttgga agggctaatt cactcccaaa gaagacaaga 5581 tatccttgat ctgtggatct accacacaca aggctacttc cctgattagc agaactacac 5641 accagggcca ggggtcagat atccactgac ctttggatgg tgctacaagc tagtaccagt 5701 tgagccagat aaggtagaag aggccaataa aggagagaac accagcttgt tacaccctgt 5761 gagcctgcat gggatggatg acccggagag agaagtgtta gagtggaggt ttgacagccg 5821 cctagcattt catcacgtgg cccgagagct gcatccggag tacttcaaga actgctgata 5881 tcgagcttgc tacaagggac tttccgctgg ggactttcca gggaggcgtg gcctgggcgg 5941 gactggggag tggcgagccc tcagatcctg catataagca gctgcttttt gcctgtactg 6001 ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac 6061 tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt 6121 gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca 6181 gtggcgcccg aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag 6241 gactcggctt gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc 6301 aaaaattttg actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa 6361 gcgggggaga attagatcgc gatgggaaaa aattcggtta aggccagggg gaaagaaaaa 6421 atataaatta aaacatatag tatgggcaag cagggagcta gaacgattcg cagttaatcc 6481 tggcctgtta gaaacatcag aaggctgtag acaaatactg ggacagctac aaccatccct 6541 tcagacagga tcagaagaac ttagatcatt atataataca gtagcaaccc tctattgtgt 6601 gcatcaaagg atagagataa aagacaccaa ggaagcttta gacaagatag aggaagagca 6661 aaacaaaagt aagaccaccg cacagcaagc ggccggccgc tgatcttcag acctggagga 6721 ggagatatga gggacaattg gagaagtgaa ttatataaat ataaagtagt aaaaattgaa 6781 ccattaggag tagcacccac caaggcaaag agaagagtgg tgcagagaga aaaaagagca 6841 gtgggaatag gagctttgtt ccttgggttc ttgggagcag caggaagcac tatgggcgca 6901 gcgtcaatga cgctgacggt acaggccaga caattattgt ctggtatagt gcagcagcag 6961 aacaatttgc tgagggctat tgaggcgcaa cagcatctgt tgcaactcac agtctggggc 7021 atcaagcagc tccaggcaag aatcctggct gtggaaagat 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 ccagtttgga tcgataagct tgatatcgaa ttcctgcagc cccgataaaa taaaagattt 7801 tatttagtct ccagaaaaag gggggaatga aagaccccac ctgtaggttt ggcaagctag 7861 ctgcagtaac gccattttgc aaggcatgga aaaataccaa accaagaata gagaagttca 7921 gatcaagggc gggtacatga aaatagctaa cgttgggcca aacaggatat ctgcggtgag 7981 cagtttcggc cccggcccgg ggccaagaac agatggtcac cgcagtttcg gccccggccc 8041 gaggccaaga acagatggtc cccagatatg gcccaaccct cagcagtttc ttaagaccca 8101 tcagatgttt ccaggctccc ccaaggacct gaaatgaccc tgcgccttat ttgaattaac 8161 caatcagcct gcttctcgct tctgttcgcg cgcttctgct tcccgagctc tataaaagag 8221 ctcacaaccc ctcactcggc gcgccagtcc tccgacagac tgagtcgccc gggggggatc 8281 tggagctctc gagaattctc acgcgtctgc aggatatcaa gcttgcggta ccgcgggccc 8341 gggatccgcc accatggatg ctaagtcact aactgcctgg tcccggacac tggtgacctt 8401 caaggatgta tttgtggact tcaccaggga ggagtggaag ctgctggaca ctgctcagca 8461 gatcgtgtac agaaatgtga tgctggagaa ctataagaac ctggtttcct tgggttatca 8521 gcttactaag ccagatgtga tcctccggtt ggagaaggga gaagagcccg gcggttccgg 8581 cggagggtcg atggacaaga agtacagcat cggcctggcc atcggcacca actctgtggg 8641 ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa 8701 caccgaccgg cacagcatca agaagaacct gatcggcgcc ctgctgttcg acagcggaga 8761 aacagccgag gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa 8821 ccggatctgc tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt 8881 cttccacaga ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc 8941 catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca 9001 cctgagaaag aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc 9061 cctggcccac atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga 9121 caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga 9181 ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag 9241 caagagcaga cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct 9301 gttcggcaac ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga 9361 cctggccgag gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa 9421 cctgctggcc cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc 9481 cgacgccatc ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct 9541 gagcgcctct atgatcaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc 9601 tctcgtgcgg cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa 9661 cggctacgcc ggctacatcg atggcggagc cagccaggaa gagttctaca agttcatcaa 9721 gcccatcctg gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga 9781 cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg 9841 agagctgcac gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg 9901 ggaaaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag 9961 gggaaacagc agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa 10021 cttcgaggaa gtggtggaca agggcgccag cgcccagagc ttcatcgagc ggatgaccaa 10081 cttcgataag aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta 10141 cttcaccgtg tacaacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc 10201 cgccttcctg agcggcgagc agaaaaaagc catcgtggac ctgctgttca agaccaaccg 10261 gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc 10321 cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct 10381 gctgaaaatt atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga 10441 agatatcgtg ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa 10501 aacctatgcc cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac 10561 cggctggggc aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa 10621 gacaatcctg gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat 10681 ccacgacgac agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg 10741 cgatagcctg cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat 10801 cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga 10861 gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag 10921 ccgcgagaga atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa 10981 agaacacccc gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca 11041 gaatgggcgg gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga 11101 tgtggacgct atcgtgcctc agagctttct gaaggacgac tccatcgata acaaagtgct 11161 gactcggagc gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa 11221 gaagatgaag aactactggc gccagctgct gaatgccaag ctgattaccc agaggaagtt 11281 cgacaatctg accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat 11341 caagagacag ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc 11401 ccggatgaac actaagtacg acgagaacga caaactgatc cgggaagtga aagtgatcac 11461 cctgaagtcc aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga 11521 gatcaacaac taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct 11581 gatcaaaaag taccctaagc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga 11641 cgtgcggaag atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt 11701 cttctacagc aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat 11761 ccggaagcgg cctctgatcg agacaaacgg cgaaacaggc gagatcgtgt gggataaggg 11821 ccgggacttt gccaccgtgc ggaaagtgct gtctatgccc caagtgaata tcgtgaaaaa 11881 gaccgaggtg cagacaggcg gcttcagcaa agagtctatc ctgcccaaga ggaacagcga 11941 caagctgatc gccagaaaga aggactggga ccctaagaag tacggcggct tcgacagccc 12001 caccgtggcc tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact 12061 gaagagtgtg aaagagctgc tggggatcac catcatggaa agaagcagct tcgagaagaa 12121 tcccatcgac tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa 12181 gctgcctaag tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc 12241 cggcgaactg cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta 12301 cctggccagc cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct 12361 gtttgtggaa cagcacaaac actacctgga cgagatcatc gagcagatca gcgagttctc 12421 caagagagtg atcctggccg acgctaatct ggacaaggtg ctgagcgcct acaacaagca 12481 cagagacaag cctatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa 12541 tctgggagcc cctgccgcct tcaagtactt tgacaccacc atcgaccgga agaggtacac 12601 cagcaccaaa gaggtgctgg acgccaccct gatccaccag agcatcaccg gcctgtacga 12661 gacacggatc gacctgtctc agctgggagg cgacgcctat ccctatgacg tgcccgatta 12721 tgccagcctg ggcagcggct cccccaagaa aaaacgcaag gtggaagatc ctaagaaaaa 12781 gcggaaagtg gacggcattg gtagtgggag caacggcagc agcggatccg ctactaactt 12841 cagcctgctg aagcaggctg gagacgtgga ggagaaccct ggacctatgg tgagcaaggg 12901 cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg 12961 ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg 13021 cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat 13081 cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc 13141 cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga 13201 ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta 13261 caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac 13321 catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga 13381 gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac 13441 ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga 13501 catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg 13561 ccactccacc ggcggcatgg acgagctgta caagtaagcg gccgcgactc tagagtcgac 13621 ctgcaggcat gcaagcttga tatcaagctt atcgataatc aacctctgga ttacaaaatt 13681 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct 13741 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg 13801 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc 13861 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt 13921 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc 13981 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg 14041 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg 14101 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc 14161 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg 14221 atctcccttt gggccgcctc cccgcat //