| 质粒名称: | lentiCRISPRv2 neo |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 98292 |
| 存储实验室: | Brett Stringer |
|
载体骨架基本信息 |
|
| 载体名称: | lentiCRISPRv2 |
| 载体出品公司: | Feng Zhang (Addgene #52961) |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 15072 bp |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 慢病毒，CRISPR |
| 筛选标记: | 新霉素（G418） |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | P2a-neo |
| 物种来源: | -- |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | EF-1a |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | BamHI (未破坏) |
| 3' 克隆位点: | MluI (未破坏) |
| 5' 测序引物 | CCAAAGAGGTGCTGGACG |
| 3' 测序引物 | WPRE-R (CATAGCGTAAAAGGAGCAACA) |
|
文献引用方法 |
|
| 材料和方法部分: | lentiCRISPRv2 neo was a gift from Brett Stringer (Addgene plasmid # 98292 ; http://n2t.net/addgene:98292 ; RRID:Addgene_98292) |
| 参考文献部分: | A reference collection of patient-derived cell line and xenograft models of proneural, classical and mesenchymal glioblastoma. Stringer BW, Day BW, D'Souza RCJ, Jamieson PR, Ensbey KS, Bruce ZC, Lim YC, Goasdoue K, Offenhauser C, Akgul S, Allan S, Robertson T, Lucas P, Tollesson G, Campbell S, Winter C, Do H, Dobrovic A, Inglis PL, Jeffree RL, Johns TG, Boyd AW. Sci Rep. 2019 Mar 20;9(1):4902. doi: 10.1038/s41598-019-41277-z. 10.1038/s41598-019-41277-z PubMed 30894629 |
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lentiCRISPRv2neo载体基本信息
lentiCRISPRv2neo载体图谱质粒图谱和多克隆位点信息
lentiCRISPRv2neo载体简介
lentiCRISPRv2neo载体序列
LOCUS lentiCRISPRv2 neo 15072 bp ds-DNA circular SYN 01-FEB-2019 DEFINITION Variant of lentiCRISPRv2 that confers G418 resistance. ACCESSION . VERSION . KEYWORDS lentiCRISPRv2 neo SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 15072) AUTHORS Brett Stringer TITLE Brett Stringer Lab Plasmids JOURNAL Unpublished REFERENCE 2 (bases 1 to 15072) AUTHORS . TITLE Direct Submission JOURNAL Exported Feb 1, 2019 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..15072 /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 242..621 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 623..821 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 776..796 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 839..1019 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 1066..1191 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1684..1917 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." misc_feature 2444..2561 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2612..2852 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 2612..2632 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 2783..2802 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_RNA 4742..4817 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter 4879..5090 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" regulatory 5109..5118 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 5115..9218 /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL DATLIHQSITGLYETRIDLSQLGGD" CDS 9219..9266 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 9267..9290 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 9300..9356 /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 9357..10148 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="IEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPV LFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSS HLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQG LAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIAL ATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(9408..9427) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 10018..10037 /label=Neo-F /note="Neomycin resistance gene, forward primer" misc_feature 10164..10752 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(10217..10237) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(10635..10646) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 10824..11057 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" primer_bind complement(11083..11100) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 11089..11313 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 11359..11787 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(11446..11465) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 11656..11677 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(11796..11816) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 11801..12130 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 11981..12116 /label=SV40 ori /note="SV40 origin of replication" primer_bind 12043..12062 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 12178..12225 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 12244..12618 /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 12748..12869 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(12785..12804) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 12839..12858 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(12918..12934) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(12918..12934) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(12931..12953) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 12942..12958 /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(12966..12996) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 13011..13032 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(13149..13166) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(13320..13908) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(13400..13419) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(14079..14939) /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 14702..14721 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(14940..15044) /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 acgcgcgcgt 241 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc 301 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc 361 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg 421 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat 481 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc 541 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta 601 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag 661 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt 721 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa 781 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gcgcgttttg cctgtactgg 841 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact 901 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg 961 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag 1021 tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg 1081 actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca 1141 aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag 1201 cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa 1261 tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct 1321 ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt 1381 cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg 1441 catcaaagga tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa 1501 aacaaaagta agaccaccgc acagcaagcg gccgctgatc ttcagacctg gaggaggaga 1561 tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt 1621 aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg 1681 aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc 1741 aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa 1801 tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa 1861 gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg 1921 gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg 1981 gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga 2041 aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga 2101 aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa 2161 cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg 2221 tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc 2281 attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga 2341 agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact 2401 gcgtgcgcca attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg 2461 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa 2521 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca 2581 gcagagatcc agtttggtta attaaggtac cgagggccta tttcccatga ttccttcata 2641 tttgcatata cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac 2701 aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt 2761 tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga 2821 tttcttggct ttatatatct tgtggaaagg acgaaacacc ggagacggtt gtaaatgagc 2881 acacaaaata cacatgctaa aatattatat tctatgacct ttataaaatc aaccaaaatc 2941 ttctttttaa taactttagt atcaataatt agaattttta tgttcctttt tgcaaacttt 3001 taataaaaat gagcaaaata aaaaaacgct agttttagta actcgcgttg ttttcttcac 3061 ctttaataat agctactcca ccacttgttc ctaagcggtc agctcctgct tcaatcattt 3121 tttgagcatc ttcaaatgtt ctaactccac cagctgcttt aactaaagca ttgtctttaa 3181 caactgactt cattagttta acatcttcaa atgttgcacc tgattttgaa aatcctgttg 3241 atgttttaac aaattctaat ccagcttcaa cagctatttc acaagctttc atgatttctt 3301 cttttgttaa taaacaattt tccataatac atttaacaac atgtgatcca gctgcttttt 3361 ttacagcttt catgtcttct aaaactaatt cataattttt gtcttttaat gcaccaatat 3421 ttaataccat atcaatttct gttgcaccat ctttaattgc ttcagaaact tcgaatgctt 3481 ttgtagctgt tgtgcatgca cctagaggaa aacctacaac atttgttatt cctacatttg 3541 tgccttttaa taattcttta caatagcttg ttcaatatga attaacacaa actgttgcaa 3601 aatcaaattc aattgcttca tcacataatt gtttaatttc agctttcgta gcatcttgtt 3661 ttaataatgt gtgatctata tatttgttta gtttcatttt ttctcctata tattcatttt 3721 taattttaat tctttaataa tttcgtctac tttaacttta gcgttttgaa cagattcacc 3781 aacacctata aaataaattt ttagtttagg ttcagttcca cttgggcgaa cagcaaatca 3841 tgacttatct tctaaataaa attttagtaa gtcttgtcct ggcatattat acattccatc 3901 gatgtagtct tcaacattaa caactttaag tccagcaatt tgagttaagg gtgttgctct 3961 caatgatttc attaatggtt caatttttaa tttcttttct tctggtttaa aattcaagtt 4021 taaagtgaaa gtgtaatatg cacccatttc tttaaataaa tcttctaaat agtctactaa 4081 tgttttattt tgttttttat aaaatcaagc agcctctgct attaatatag aagcttgtat 4141 tccatcttta tctctagctg agtcatcaat tacatatcca taactttctt cataagcaaa 4201 aacaaaattt aatccgttat cttcttcttt agcaatttct ctacccattc atttaaatcc 4261 agttaaagtt tttacaatat taactccata tttttcatga gcgattctat cacccaaatc 4321 acttgttaca aaacttgaat atagagccgg attttttgga atgctattta agcgttttag 4381 atttgataat tttcaatcaa ttaaaattgg tcctgtttga tttccatcta atcttacaaa 4441 atgaccatca tgttttattg ccattccaaa tctgtcagca tctgggtcat tcataataat 4501 aatatctgca tcatgtttaa taccatattc aagcggtatt tttcatgcag gatcaaattc 4561 tggatttgga tttacaacat ttttaaatgt ttcatcttca aatgcatgct cttcaacctc 4621 aataacgtta tatcctgatt cacgtaatat ttttggggta aatttagttc ctgttccatt 4681 aactgcgcta aaaataattt ttaaatcttt tttagcttct tgctcttttt tgtacgtctc 4741 tgttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag 4801 tggcaccgag tcggtgcttt tttgaattcg ctagctaggt cttgaaagga gtgggaattg 4861 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg 4921 gaggggtcgg caattgatcc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg 4981 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag 5041 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gaccggttct 5101 agagcgctgc caccatggac aagaagtaca gcatcggcct ggacatcggc accaactctg 5161 tgggctgggc cgtgatcacc gacgagtaca aggtgcccag caagaaattc aaggtgctgg 5221 gcaacaccga ccggcacagc atcaagaaga acctgatcgg agccctgctg ttcgacagcg 5281 gcgaaacagc cgaggccacc cggctgaaga gaaccgccag aagaagatac accagacgga 5341 agaaccggat ctgctatctg caagagatct tcagcaacga gatggccaag gtggacgaca 5401 gcttcttcca cagactggaa gagtccttcc tggtggaaga ggataagaag cacgagcggc 5461 accccatctt cggcaacatc gtggacgagg tggcctacca cgagaagtac cccaccatct 5521 accacctgag aaagaaactg gtggacagca ccgacaaggc cgacctgcgg ctgatctatc 5581 tggccctggc ccacatgatc aagttccggg gccacttcct gatcgagggc gacctgaacc 5641 ccgacaacag cgacgtggac aagctgttca tccagctggt gcagacctac aaccagctgt 5701 tcgaggaaaa ccccatcaac gccagcggcg tggacgccaa ggccatcctg tctgccagac 5761 tgagcaagag cagacggctg gaaaatctga tcgcccagct gcccggcgag aagaagaatg 5821 gcctgttcgg aaacctgatt gccctgagcc tgggcctgac ccccaacttc aagagcaact 5881 tcgacctggc cgaggatgcc aaactgcagc tgagcaagga cacctacgac gacgacctgg 5941 acaacctgct ggcccagatc ggcgaccagt acgccgacct gtttctggcc gccaagaacc 6001 tgtccgacgc catcctgctg agcgacatcc tgagagtgaa caccgagatc accaaggccc 6061 ccctgagcgc ctctatgatc aagagatacg acgagcacca ccaggacctg accctgctga 6121 aagctctcgt gcggcagcag ctgcctgaga agtacaaaga gattttcttc gaccagagca 6181 agaacggcta cgccggctac attgacggcg gagccagcca ggaagagttc tacaagttca 6241 tcaagcccat cctggaaaag atggacggca ccgaggaact gctcgtgaag ctgaacagag 6301 aggacctgct gcggaagcag cggaccttcg acaacggcag catcccccac cagatccacc 6361 tgggagagct gcacgccatt ctgcggcggc aggaagattt ttacccattc ctgaaggaca 6421 accgggaaaa gatcgagaag atcctgacct tccgcatccc ctactacgtg ggccctctgg 6481 ccaggggaaa cagcagattc gcctggatga ccagaaagag cgaggaaacc atcaccccct 6541 ggaacttcga ggaagtggtg gacaagggcg cttccgccca gagcttcatc gagcggatga 6601 ccaacttcga taagaacctg cccaacgaga aggtgctgcc caagcacagc ctgctgtacg 6661 agtacttcac cgtgtataac gagctgacca aagtgaaata cgtgaccgag ggaatgagaa 6721 agcccgcctt cctgagcggc gagcagaaaa aggccatcgt ggacctgctg ttcaagacca 6781 accggaaagt gaccgtgaag cagctgaaag aggactactt caagaaaatc gagtgcttcg 6841 actccgtgga aatctccggc gtggaagatc ggttcaacgc ctccctgggc acataccacg 6901 atctgctgaa aattatcaag gacaaggact tcctggacaa tgaggaaaac gaggacattc 6961 tggaagatat cgtgctgacc ctgacactgt ttgaggacag agagatgatc gaggaacggc 7021 tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag cggcggagat 7081 acaccggctg gggcaggctg agccggaagc tgatcaacgg catccgggac aagcagtccg 7141 gcaagacaat cctggatttc ctgaagtccg acggcttcgc caacagaaac ttcatgcagc 7201 tgatccacga cgacagcctg acctttaaag aggacatcca gaaagcccag gtgtccggcc 7261 agggcgatag cctgcacgag cacattgcca atctggccgg cagccccgcc attaagaagg 7321 gcatcctgca gacagtgaag gtggtggacg agctcgtgaa agtgatgggc cggcacaagc 7381 ccgagaacat cgtgatcgaa atggccagag agaaccagac cacccagaag ggacagaaga 7441 acagccgcga gagaatgaag cggatcgaag agggcatcaa agagctgggc agccagatcc 7501 tgaaagaaca ccccgtggaa aacacccagc tgcagaacga gaagctgtac ctgtactacc 7561 tgcagaatgg gcgggatatg tacgtggacc aggaactgga catcaaccgg ctgtccgact 7621 acgatgtgga ccatatcgtg cctcagagct ttctgaagga cgactccatc gacaacaagg 7681 tgctgaccag aagcgacaag aaccggggca agagcgacaa cgtgccctcc gaagaggtcg 7741 tgaagaagat gaagaactac tggcggcagc tgctgaacgc caagctgatt acccagagaa 7801 agttcgacaa tctgaccaag gccgagagag gcggcctgag cgaactggat aaggccggct 7861 tcatcaagag acagctggtg gaaacccggc agatcacaaa gcacgtggca cagatcctgg 7921 actcccggat gaacactaag tacgacgaga atgacaagct gatccgggaa gtgaaagtga 7981 tcaccctgaa gtccaagctg gtgtccgatt tccggaagga tttccagttt tacaaagtgc 8041 gcgagatcaa caactaccac cacgcccacg acgcctacct gaacgccgtc gtgggaaccg 8101 ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt 8161 acgacgtgcg gaagatgatc gccaagagcg agcaggaaat cggcaaggct accgccaagt 8221 acttcttcta cagcaacatc atgaactttt tcaagaccga gattaccctg gccaacggcg 8281 agatccggaa gcggcctctg atcgagacaa acggcgaaac cggggagatc gtgtgggata 8341 agggccggga ttttgccacc gtgcggaaag tgctgagcat gccccaagtg aatatcgtga 8401 aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc aagaggaaca 8461 gcgataagct gatcgccaga aagaaggact gggaccctaa gaagtacggc ggcttcgaca 8521 gccccaccgt ggcctattct gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga 8581 aactgaagag tgtgaaagag ctgctgggga tcaccatcat ggaaagaagc agcttcgaga 8641 agaatcccat cgactttctg gaagccaagg gctacaaaga agtgaaaaag gacctgatca 8701 tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg ccggaagaga atgctggcct 8761 ctgccggcga actgcagaag ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc 8821 tgtacctggc cagccactat gagaagctga agggctcccc cgaggataat gagcagaaac 8881 agctgtttgt ggaacagcac aagcactacc tggacgagat catcgagcag atcagcgagt 8941 tctccaagag agtgatcctg gccgacgcta atctggacaa agtgctgtcc gcctacaaca 9001 agcaccggga taagcccatc agagagcagg ccgagaatat catccacctg tttaccctga 9061 ccaatctggg agcccctgcc gccttcaagt actttgacac caccatcgac cggaagaggt 9121 acaccagcac caaagaggtg ctggacgcca ccctgatcca ccagagcatc accggcctgt 9181 acgagacacg gatcgacctg tctcagctgg gaggcgacaa gcgacctgcc gccacaaaga 9241 aggctggaca ggctaagaag aagaaagatt acaaagacga tgacgataag ggatccggcg 9301 caacaaactt ctctctgctg aaacaagccg gagatgtcga agagaatcct ggaccgattg 9361 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg 9421 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg 9481 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg 9541 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg 9601 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc 9661 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc 9721 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc 9781 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc 9841 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg 9901 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct 9961 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt 10021 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc 10081 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt 10141 tcttctgaac gcgttaagtc gacaatcaac ctctggatta caaaatttgt gaaagattga 10201 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt 10261 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt 10321 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg 10381 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg 10441 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc 10501 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat 10561 catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct 10621 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg 10681 ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg 10741 ccgcctcccc gcgtcgactt taagaccaat gacttacaag gcagctgtag atcttagcca 10801 ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa gacaagatct 10861 gctttttgct tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg 10921 ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt 10981 gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt 11041 gtggaaaatc tctagcaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta 11101 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca 11161 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc 11221 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata 11281 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg 11341 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg 11401 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct 11461 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc 11521 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg 11581 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt 11641 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg 11701 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc 11761 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg 11821 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc 11881 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct 11941 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc 12001 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga 12061 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg 12121 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc agcacgtgtt 12181 gacaattaat catcggcata gtatatcggc atagtataat acgacaaggt gaggaactaa 12241 accatggcca agttgaccag tgccgttccg gtgctcaccg cgcgcgacgt cgccggagcg 12301 gtcgagttct ggaccgaccg gctcgggttc tcccgggact tcgtggagga cgacttcgcc 12361 ggtgtggtcc gggacgacgt gaccctgttc atcagcgcgg tccaggacca ggtggtgccg 12421 gacaacaccc tggcctgggt gtgggtgcgc ggcctggacg agctgtacgc cgagtggtcg 12481 gaggtcgtgt ccacgaactt ccgggacgcc tccgggccgg ccatgaccga gatcggcgag 12541 cagccgtggg ggcgggagtt cgccctgcgc gacccggccg gcaactgcgt gcacttcgtg 12601 gccgaggagc aggactgaca cgtgctacga gatttcgatt ccaccgccgc cttctatgaa 12661 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 12721 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa 12781 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt 12841 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag 12901 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt 12961 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc 13021 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc 13081 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct 13141 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 13201 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 13261 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 13321 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 13381 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 13441 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 13501 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 13561 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 13621 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 13681 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 13741 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc 13801 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 13861 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 13921 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 13981 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 14041 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 14101 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg 14161 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 14221 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag 14281 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa 14341 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc 14401 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca 14461 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 14521 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat 14581 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 14641 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg 14701 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 14761 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt 14821 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca 14881 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 14941 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac 15001 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa 15061 gtgccacctg ac //
