LOCUS Exported 14873 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Lentiviral plasmid for expression of gRNA and dCas9 in mammalian cells; derivative of lentiCRISPR v2. ACCESSION . VERSION . KEYWORDS lentiCRISPR v2-dCas9 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 14873) AUTHORS Babaei M, Liu Y, Wuerzberger-Davis SM, McCaslin EZ, DiRusso CJ, Yeo AT, Kagermazova L, Miyamoto S, Gilmore TD TITLE CRISPR/Cas9-based editing of a sensitive transcriptional regulatory element to achieve cell type-specific knockdown of the NEMO scaffold protein. JOURNAL PLoS One. 2019 Sep 25;14(9):e0222588. doi: 10.1371/journal.pone.0222588. eCollection 2019. PUBMED 31553754 REFERENCE 2 (bases 1 to 14873) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..14873 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 316..433 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 484..724 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 484..504 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 655..674 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_RNA 2614..2689 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter 2751..2962 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" regulatory 2981..2990 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2987..7090 /codon_start=1 /product="Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system" /label=Cas9 /note="generates RNA-guided double strand breaks in DNA" /translation="MDKKYSIGLGIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL DATLIHQSITGLYETRIDLSQLGGD" CDS 7091..7138 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 7139..7162 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 7172..7228 /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 7229..7825 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind 7722..7742 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 7841..8429 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(7894..7914) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(8312..8323) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 8501..8734 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" primer_bind complement(8760..8777) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 8766..8990 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 9036..9464 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9123..9142) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 9333..9354 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(9473..9493) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 9478..9807 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 9658..9793 /label=SV40 ori /note="SV40 origin of replication" primer_bind 9720..9739 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 9855..9902 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 9921..10295 /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 10425..10546 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(10462..10481) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 10516..10535 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(10595..10611) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(10595..10611) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(10608..10630) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 10619..10635 /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(10643..10673) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 10688..10709 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(10826..10843) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(10997..11585) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(11077..11096) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(11756..12616) /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 12379..12398 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(12617..12721) /gene="bla" /label=AmpR promoter primer_bind complement(12796..12815) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 12987..13366 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 13368..13566 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 13521..13541 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 13584..13764 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 13811..13936 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 14429..14662 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 14847..18 /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" ORIGIN 1 gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg gtatataaaa 61 ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc tgtactttct 121 atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc 181 ccgaggggac ccgacaggcc cgaaggaata gaagaagaag gtggagagag agacagagac 241 agatccattc gattagtgaa cggatcggca ctgcgtgcgc caattctgca gacaaatggc 301 agtattcatc cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag 361 aatagtagac ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa 421 aattcaaaat tttcgggttt attacaggga cagcagagat ccagtttggt taattaaggt 481 accgagggcc tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga 541 gagataatta gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt 601 agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat 661 catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa 721 ggacgaaaca ccggagacgg ttgtaaatga gcacacaaaa tacacatgct aaaatattat 781 attctatgac ctttataaaa tcaaccaaaa tcttcttttt aataacttta gtatcaataa 841 ttagaatttt tatgttcctt tttgcaaact tttaataaaa atgagcaaaa taaaaaaacg 901 ctagttttag taactcgcgt tgttttcttc acctttaata atagctactc caccacttgt 961 tcctaagcgg tcagctcctg cttcaatcat tttttgagca tcttcaaatg ttctaactcc 1021 accagctgct ttaactaaag cattgtcttt aacaactgac ttcattagtt taacatcttc 1081 aaatgttgca cctgattttg aaaatcctgt tgatgtttta acaaattcta atccagcttc 1141 aacagctatt tcacaagctt tcatgatttc ttcttttgtt aataaacaat tttccataat 1201 acatttaaca acatgtgatc cagctgcttt ttttacagct ttcatgtctt ctaaaactaa 1261 ttcataattt ttgtctttta atgcaccaat atttaatacc atatcaattt ctgttgcacc 1321 atctttaatt gcttcagaaa cttcgaatgc ttttgtagct gttgtgcatg cacctagagg 1381 aaaacctaca acatttgtta ttcctacatt tgtgcctttt aataattctt tacaatagct 1441 tgttcaatat gaattaacac aaactgttgc aaaatcaaat tcaattgctt catcacataa 1501 ttgtttaatt tcagctttcg tagcatcttg ttttaataat gtgtgatcta tatatttgtt 1561 tagtttcatt ttttctccta tatattcatt tttaatttta attctttaat aatttcgtct 1621 actttaactt tagcgttttg aacagattca ccaacaccta taaaataaat ttttagttta 1681 ggttcagttc cacttgggcg aacagcaaat catgacttat cttctaaata aaattttagt 1741 aagtcttgtc ctggcatatt atacattcca tcgatgtagt cttcaacatt aacaacttta 1801 agtccagcaa tttgagttaa gggtgttgct ctcaatgatt tcattaatgg ttcaattttt 1861 aatttctttt cttctggttt aaaattcaag tttaaagtga aagtgtaata tgcacccatt 1921 tctttaaata aatcttctaa atagtctact aatgttttat tttgtttttt ataaaatcaa 1981 gcagcctctg ctattaatat agaagcttgt attccatctt tatctctagc tgagtcatca 2041 attacatatc cataactttc ttcataagca aaaacaaaat ttaatccgtt atcttcttct 2101 ttagcaattt ctctacccat tcatttaaat ccagttaaag tttttacaat attaactcca 2161 tatttttcat gagcgattct atcacccaaa tcacttgtta caaaacttga atatagagcc 2221 ggattttttg gaatgctatt taagcgtttt agatttgata attttcaatc aattaaaatt 2281 ggtcctgttt gatttccatc taatcttaca aaatgaccat catgttttat tgccattcca 2341 aatctgtcag catctgggtc attcataata ataatatctg catcatgttt aataccatat 2401 tcaagcggta tttttcatgc aggatcaaat tctggatttg gatttacaac atttttaaat 2461 gtttcatctt caaatgcatg ctcttcaacc tcaataacgt tatatcctga ttcacgtaat 2521 atttttgggg taaatttagt tcctgttcca ttaactgcgc taaaaataat ttttaaatct 2581 tttttagctt cttgctcttt tttgtacgtc tctgttttag agctagaaat agcaagttaa 2641 aataaggcta gtccgttatc aacttgaaaa agtggcaccg agtcggtgct tttttgaatt 2701 cgctagctag gtcttgaaag gagtgggaat tggctccggt gcccgtcagt gggcagagcg 2761 cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgat ccggtgccta 2821 gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc 2881 cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa 2941 cgggtttgcc gccagaacac aggaccggtt ctagagcgct gccaccatgg acaagaagta 3001 cagcatcggc ctgggcatcg gcaccaactc tgtgggctgg gccgtgatca ccgacgagta 3061 caaggtgccc agcaagaaat tcaaggtgct gggcaacacc gaccggcaca gcatcaagaa 3121 gaacctgatc ggagccctgc tgttcgacag cggcgaaaca gccgaggcca cccggctgaa 3181 gagaaccgcc agaagaagat acaccagacg gaagaaccgg atctgctatc tgcaagagat 3241 cttcagcaac gagatggcca aggtggacga cagcttcttc cacagactgg aagagtcctt 3301 cctggtggaa gaggataaga agcacgagcg gcaccccatc ttcggcaaca tcgtggacga 3361 ggtggcctac cacgagaagt accccaccat ctaccacctg agaaagaaac tggtggacag 3421 caccgacaag gccgacctgc ggctgatcta tctggccctg gcccacatga tcaagttccg 3481 gggccacttc ctgatcgagg gcgacctgaa ccccgacaac agcgacgtgg acaagctgtt 3541 catccagctg gtgcagacct acaaccagct gttcgaggaa aaccccatca acgccagcgg 3601 cgtggacgcc aaggccatcc tgtctgccag actgagcaag agcagacggc tggaaaatct 3661 gatcgcccag ctgcccggcg agaagaagaa tggcctgttc ggaaacctga ttgccctgag 3721 cctgggcctg acccccaact tcaagagcaa cttcgacctg gccgaggatg ccaaactgca 3781 gctgagcaag gacacctacg acgacgacct ggacaacctg ctggcccaga tcggcgacca 3841 gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac gccatcctgc tgagcgacat 3901 cctgagagtg aacaccgaga tcaccaaggc ccccctgagc gcctctatga tcaagagata 3961 cgacgagcac caccaggacc tgaccctgct gaaagctctc gtgcggcagc agctgcctga 4021 gaagtacaaa gagattttct tcgaccagag caagaacggc tacgccggct acattgacgg 4081 cggagccagc caggaagagt tctacaagtt catcaagccc atcctggaaa agatggacgg 4141 caccgaggaa ctgctcgtga agctgaacag agaggacctg ctgcggaagc agcggacctt 4201 cgacaacggc agcatccccc accagatcca cctgggagag ctgcacgcca ttctgcggcg 4261 gcaggaagat ttttacccat tcctgaagga caaccgggaa aagatcgaga agatcctgac 4321 cttccgcatc ccctactacg tgggccctct ggccagggga aacagcagat tcgcctggat 4381 gaccagaaag agcgaggaaa ccatcacccc ctggaacttc gaggaagtgg tggacaaggg 4441 cgcttccgcc cagagcttca tcgagcggat gaccaacttc gataagaacc tgcccaacga 4501 gaaggtgctg cccaagcaca gcctgctgta cgagtacttc accgtgtata acgagctgac 4561 caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc ttcctgagcg gcgagcagaa 4621 aaaggccatc gtggacctgc tgttcaagac caaccggaaa gtgaccgtga agcagctgaa 4681 agaggactac ttcaagaaaa tcgagtgctt cgactccgtg gaaatctccg gcgtggaaga 4741 tcggttcaac gcctccctgg gcacatacca cgatctgctg aaaattatca aggacaagga 4801 cttcctggac aatgaggaaa acgaggacat tctggaagat atcgtgctga ccctgacact 4861 gtttgaggac agagagatga tcgaggaacg gctgaaaacc tatgcccacc tgttcgacga 4921 caaagtgatg aagcagctga agcggcggag atacaccggc tggggcaggc tgagccggaa 4981 gctgatcaac ggcatccggg acaagcagtc cggcaagaca atcctggatt tcctgaagtc 5041 cgacggcttc gccaacagaa acttcatgca gctgatccac gacgacagcc tgacctttaa 5101 agaggacatc cagaaagccc aggtgtccgg ccagggcgat agcctgcacg agcacattgc 5161 caatctggcc ggcagccccg ccattaagaa gggcatcctg cagacagtga aggtggtgga 5221 cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac atcgtgatcg aaatggccag 5281 agagaaccag accacccaga agggacagaa gaacagccgc gagagaatga agcggatcga 5341 agagggcatc aaagagctgg gcagccagat cctgaaagaa caccccgtgg aaaacaccca 5401 gctgcagaac gagaagctgt acctgtacta cctgcagaat gggcgggata tgtacgtgga 5461 ccaggaactg gacatcaacc ggctgtccga ctacgatgtg gacgccatcg tgcctcagag 5521 ctttctgaag gacgactcca tcgacaacaa ggtgctgacc agaagcgaca agaaccgggg 5581 caagagcgac aacgtgccct ccgaagaggt cgtgaagaag atgaagaact actggcggca 5641 gctgctgaac gccaagctga ttacccagag aaagttcgac aatctgacca aggccgagag 5701 aggcggcctg agcgaactgg ataaggccgg cttcatcaag agacagctgg tggaaacccg 5761 gcagatcaca aagcacgtgg cacagatcct ggactcccgg atgaacacta agtacgacga 5821 gaatgacaag ctgatccggg aagtgaaagt gatcaccctg aagtccaagc tggtgtccga 5881 tttccggaag gatttccagt tttacaaagt gcgcgagatc aacaactacc accacgccca 5941 cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc aaaaagtacc ctaagctgga 6001 aagcgagttc gtgtacggcg actacaaggt gtacgacgtg cggaagatga tcgccaagag 6061 cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc tacagcaaca tcatgaactt 6121 tttcaagacc gagattaccc tggccaacgg cgagatccgg aagcggcctc tgatcgagac 6181 aaacggcgaa accggggaga tcgtgtggga taagggccgg gattttgcca ccgtgcggaa 6241 agtgctgagc atgccccaag tgaatatcgt gaaaaagacc gaggtgcaga caggcggctt 6301 cagcaaagag tctatcctgc ccaagaggaa cagcgataag ctgatcgcca gaaagaagga 6361 ctgggaccct aagaagtacg gcggcttcga cagccccacc gtggcctatt ctgtgctggt 6421 ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag agtgtgaaag agctgctggg 6481 gatcaccatc atggaaagaa gcagcttcga gaagaatccc atcgactttc tggaagccaa 6541 gggctacaaa gaagtgaaaa aggacctgat catcaagctg cctaagtact ccctgttcga 6601 gctggaaaac ggccggaaga gaatgctggc ctctgccggc gaactgcaga agggaaacga 6661 actggccctg ccctccaaat atgtgaactt cctgtacctg gccagccact atgagaagct 6721 gaagggctcc cccgaggata atgagcagaa acagctgttt gtggaacagc acaagcacta 6781 cctggacgag atcatcgagc agatcagcga gttctccaag agagtgatcc tggccgacgc 6841 taatctggac aaagtgctgt ccgcctacaa caagcaccgg gataagccca tcagagagca 6901 ggccgagaat atcatccacc tgtttaccct gaccaatctg ggagcccctg ccgccttcaa 6961 gtactttgac accaccatcg accggaagag gtacaccagc accaaagagg tgctggacgc 7021 caccctgatc caccagagca tcaccggcct gtacgagaca cggatcgacc tgtctcagct 7081 gggaggcgac aagcgacctg ccgccacaaa gaaggctgga caggctaaga agaagaaaga 7141 ttacaaagac gatgacgata agggatccgg cgcaacaaac ttctctctgc tgaaacaagc 7201 cggagatgtc gaagagaatc ctggaccgac cgagtacaag cccacggtgc gcctcgccac 7261 ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc 7321 cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact 7381 cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc 7441 ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg 7501 cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct 7561 cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg gagtctcgcc 7621 cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga 7681 gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga 7741 gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg 7801 catgacccgc aagcccggtg cctgaacgcg ttaagtcgac aatcaacctc tggattacaa 7861 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata 7921 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc 7981 cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg 8041 tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac 8101 ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat 8161 cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt 8221 ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat 8281 tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc 8341 ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag 8401 tcggatctcc ctttgggccg cctccccgcg tcgactttaa gaccaatgac ttacaaggca 8461 gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc 8521 caacgaagac aagatctgct ttttgcttgt actgggtctc tctggttaga ccagatctga 8581 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct 8641 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc 8701 agaccctttt agtcagtgtg gaaaatctct agcagggccc gtttaaaccc gctgatcagc 8761 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt 8821 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca 8881 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga 8941 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc 9001 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag 9061 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc 9121 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc 9181 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa 9241 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg 9301 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac 9361 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta 9421 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg 9481 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg 9541 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt 9601 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc 9661 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt 9721 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc 9781 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga 9841 tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata gtataatacg 9901 acaaggtgag gaactaaacc atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc 9961 gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc cgggacttcg 10021 tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc agcgcggtcc 10081 aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc 10141 tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca 10201 tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac ccggccggca 10261 actgcgtgca cttcgtggcc gaggagcagg actgacacgt gctacgagat ttcgattcca 10321 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga 10381 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag 10441 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt 10501 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac 10561 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt 10621 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg 10681 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt 10741 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt 10801 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc 10861 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg 10921 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg 10981 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac 11041 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg 11101 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct 11161 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg 11221 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct 11281 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac 11341 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt 11401 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc 11461 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca 11521 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat 11581 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac 11641 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt 11701 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc 11761 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg 11821 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg 11881 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc 11941 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta 12001 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg 12061 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct 12121 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta 12181 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg 12241 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga 12301 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt 12361 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca 12421 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt 12481 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt 12541 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga 12601 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt 12661 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc 12721 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct 12781 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc 12841 tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca 12901 aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg 12961 cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc 13021 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt 13081 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta 13141 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg 13201 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga 13261 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt 13321 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg 13381 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc 13441 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg 13501 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat 13561 aagcagcgcg ttttgcctgt actgggtctc tctggttaga ccagatctga gcctgggagc 13621 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc 13681 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt 13741 agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag cgaaagggaa 13801 accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga 13861 ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga aggagagaga 13921 tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg 13981 gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg caagcaggga 14041 gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct gtagacaaat 14101 actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat cattatataa 14161 tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca ccaaggaagc 14221 tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc aagcggccgc 14281 tgatcttcag acctggagga ggagatatga gggacaattg gagaagtgaa ttatataaat 14341 ataaagtagt aaaaattgaa ccattaggag tagcacccac caaggcaaag agaagagtgg 14401 tgcagagaga aaaaagagca gtgggaatag gagctttgtt ccttgggttc ttgggagcag 14461 caggaagcac tatgggcgca gcgtcaatga cgctgacggt acaggccaga caattattgt 14521 ctggtatagt gcagcagcag aacaatttgc tgagggctat tgaggcgcaa cagcatctgt 14581 tgcaactcac agtctggggc atcaagcagc tccaggcaag aatcctggct gtggaaagat 14641 acctaaagga tcaacagctc ctggggattt ggggttgctc tggaaaactc atttgcacca 14701 ctgctgtgcc ttggaatgct agttggagta ataaatctct ggaacagatt tggaatcaca 14761 cgacctggat ggagtgggac agagaaatta acaattacac aagcttaata cactccttaa 14821 ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga attattggaa tta //