质粒名称: | pCMV-MMLVgag-3xNES-ABE8e |
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目录编号: | 181751 |
存储实验室: | David Liu |
载体骨架基本信息 |
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载体名称: | pCDNA3-like |
空载体大小: | 4757 bp |
完整质粒大小: | 11365bp |
载体类型: | 哺乳细胞表达 |
载体抗性: | Ampicillin, 100 μg/mL |
生长温度: | 37 ℃ |
宿主菌: | DH5a |
拷贝数: | 高拷贝 |
插入基因信息 |
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插入基因名称: | MMLVgag-3xNES-ABE8e |
物种来源: | H. sapiens (human), M. musculus (mouse) |
插入大小: | 6609 bp |
启动子: | CMV |
标签/融合蛋白: | Flag (N terminal on insert) |
克隆方法: | Ligation Independent Cloning |
5’测序引物 | ACCCTGGTCATCATCCTG |
3’测序引物 | CAGGAAACAGCTATGAC; CCTTCCGAGTGAGAGAC |
文献引用方法 |
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材料和方法部分: | pCMV-MMLVgag-3xNES-ABE8e was a gift from David Liu (Addgene plasmid # 181751 ; http://n2t.net/addgene:181751 ; RRID:Addgene_181751) |
参考文献部分: | Engineered virus-like particles for efficient in vivo delivery of therapeutic proteins. Banskota S, Raguram A, Suh S, Du SW, Davis JR, Choi EH, Wang X, Nielsen SC, Newby GA, Randolph PB, Osborn MJ, Musunuru K, Palczewski K, Liu DR. Cell. 2022 Jan 7. pii: S0092-8674(21)01484-7. doi: 10.1016/j.cell.2021.12.021. 10.1016/j.cell.2021.12.021 PubMed 35 |
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载体基本信息
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载体简介
载体序列
LOCUS pCMV-MMLVgag-3xNES-ABE8e 11365 bp ds-DNA circular SYN 08-FEB-2022 DEFINITION MMLV-Gag fused to 3xNES-ABE8e with TSTLLMENSS cleavage site between Gag-NES and ABE. For production of virus like particles with base-editor RNP cargo.. ACCESSION . VERSION . KEYWORDS pCMV-MMLVgag-3xNES-ABE8e SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 11365) AUTHORS Banskota S, Raguram A, Suh S, Du SW, Davis JR, Choi EH, Wang X, Nielsen SC, Newby GA, Randolph PB, Osborn MJ, Musunuru K, Palczewski K, Liu DR TITLE Engineered virus-like particles for efficient in vivo delivery of therapeutic proteins. JOURNAL Cell. 2022 Jan 7. pii: S0092-8674(21)01484-7. doi: 10.1016/j.cell.2021.12.021. PUBMED 35021064 REFERENCE 2 (bases 1 to 11365) AUTHORS . TITLE Direct Submission JOURNAL Exported Feb 8, 2022 from SnapGene Server 1.1.58 http://www.biofeng.com FEATURES Location/Qualifiers source 1..11365 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 4..420 /label=gag (truncated) /note="truncated Moloney murine leukemia virus (MMLV) gag gene lacking the start codon" primer_bind 350..372 /label=pLXSN 5' /note="Murine stem cell virus, forward primer. Also called MSCV" primer_bind 388..404 /label=pBABE 5' /note="Psi packaging signal, forward primer for pBABE vectors" CDS 1633..1659 /codon_start=1 /product="nuclear export signal from the HIV Rev protein (Fischer et al., 1995)" /label=NES /translation="LPPLERLTL" CDS 1672..1698 /codon_start=1 /product="nuclear export signal from the HIV Rev protein (Fischer et al., 1995)" /label=NES /translation="LPPLERLTL" CDS 1711..1737 /codon_start=1 /product="nuclear export signal from the HIV Rev protein (Fischer et al., 1995)" /label=NES /translation="LPPLERLTL" CDS 1771..1794 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1828..1848 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 2443..6543 /codon_start=1 /product="nickase mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system" /label=Cas9(D10A) /note="generates RNA-guided single strand nicks in DNA due to the D10A mutation in the RuvC catalytic domain" /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD ATLIHQSITGLYETRIDLSQLGGD" CDS 6586..6606 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" primer_bind complement(6612..6631) /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" polyA_signal 6677..6732 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(6731..6750) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(7090..7106) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(7090..7106) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(7103..7125) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 7114..7130 /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(7138..7168) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 7183..7204 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(7321..7338) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(7492..8080) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(7572..7591) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(8251..9111) /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 8874..8893 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(9112..9216) /gene="bla" /label=AmpR promoter rep_origin complement(9242..9697) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9379..9400) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 9591..9610 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 9824..9846 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 9838..9855 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 9839..9855 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 9865..9884 /label=T7 /note="T7 promoter, forward primer" promoter 9865..9883 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" enhancer 10004..10383 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 10384..10587 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 10538..10558 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 10584..10608 /label=LNCX /note="Human CMV promoter, forward primer" intron 10699..11271 /label=-beta-globin intron /note="intron from rabbit beta-globin gene" ORIGIN 1 atgggccaga ctgttaccac tcccttaagt ttgaccttag gtcactggaa agatgtcgag 61 cggatcgctc acaaccagtc ggtagatgtc aagaagagac gttgggttac cttctgctct 121 gcagaatggc caacctttaa cgtcggatgg ccgcgagacg gcacctttaa ccgagacctc 181 atcacccagg ttaagatcaa ggtcttttca cctggcccgc atggacaccc agaccaggtc 241 ccctacatcg tgacctggga agccttggct tttgaccccc ctccctgggt caagcccttt 301 gtacacccta agcctccgcc tcctcttcct ccatccgccc cgtctctccc ccttgaacct 361 cctcgttcga ccccgcctcg atcctccctt tatccagccc tcactccttc tctaggcgcc 421 aaacctaaac ctcaagttct ttctgacagt ggggggccgc tcatcgacct acttacagaa 481 gaccccccgc cttataggga cccaagacca cccccttccg acagggacgg aaatggtgga 541 gaagcgaccc ctgcgggaga ggcaccggac ccctccccaa tggcatctcg cctacgtggg 601 agacgggagc cccctgtggc cgactccact acctcgcagg cattccccct ccgcgcagga 661 ggaaacggac agcttcaata ctggccgttc tcctcttctg acctttacaa ctggaaaaat 721 aataaccctt ctttttctga agatccaggt aaactgacag ctctgatcga gtctgtcctc 781 atcacccatc agcccacctg ggacgactgt cagcagctgt tggggactct gctgaccgga 841 gaagaaaaac aacgggtgct cttagaggct agaaaggcgg tgcggggcga tgatgggcgc 901 cccactcaac tgcccaatga agtcgatgcc gcttttcccc tcgagcgccc agactgggat 961 tacaccaccc aggcaggtag gaaccaccta gtccactatc gccagttgct cctagcgggt 1021 ctccaaaacg cgggcagaag ccccaccaat ttggccaagg taaaaggaat aacacaaggg 1081 cccaatgagt ctccctcggc cttcctagag agacttaagg aagcctatcg caggtacact 1141 ccttatgacc ctgaggaccc agggcaagaa actaatgtgt ctatgtcttt catttggcag 1201 tctgccccag acattgggag aaagttagag aggttagaag atttaaaaaa caagacgctt 1261 ggagatttgg ttagagaggc agaaaagatc tttaataaac gagaaacccc ggaagaaaga 1321 gaggaacgta tcaggagaga aacagaggaa aaagaagaac gccgtaggac agaggatgag 1381 cagaaagaga aagaaagaga tcgtaggaga catagagaga tgagcaagct attggccact 1441 gtcgttagtg gacagaaaca ggatagacag ggaggagaac gaaggaggtc ccaactcgat 1501 cgcgaccagt gtgcctactg caaagaaaag gggcactggg ctaaagattg tcccaagaaa 1561 ccacgaggac ctcggggacc aagaccccag acctccctcc tgaccctaga tgactctggc 1621 ggctcacttc aactgcctcc acttgaaaga ctgacactgg gatcattaca attacctcct 1681 ttagaacgat taacactcgg ttcactacag cttccgcctc ttgagagatt gacattaacg 1741 tccacgctgc taatggagaa ctcgtctgga gattacaaag acgatgacga taagaaacgg 1801 acagccgacg gaagcgagtt cgagtcacca aagaagaagc ggaaagtctc tgaggtggag 1861 ttttcccacg agtactggat gagacatgcc ctgaccctgg ccaagagggc acgggatgag 1921 agggaggtgc ctgtgggagc cgtgctggtg ctgaacaata gagtgatcgg cgagggctgg 1981 aacagagcca tcggcctgca cgacccaaca gcccatgccg aaattatggc cctgagacag 2041 ggcggcctgg tcatgcagaa ctacagactg attgacgcca ccctgtacgt gacattcgag 2101 ccttgcgtga tgtgcgccgg cgccatgatc cactctagga tcggccgcgt ggtgtttggc 2161 gtgaggaact caaaaagagg cgccgcaggc tccctgatga acgtgctgaa ctaccccggc 2221 atgaatcacc gcgtcgaaat taccgaggga atcctggcag atgaatgtgc cgccctgctg 2281 tgcgatttct atcggatgcc tagacaggtg ttcaatgctc agaagaaggc ccagagctcc 2341 atcaactccg gaggatctag cggaggctcc tctggctctg agacacctgg cacaagcgag 2401 agcgcaacac ctgaaagcag cgggggcagc agcggggggt cagacaagaa gtacagcatc 2461 ggcctggcca tcggcaccaa ctctgtgggc tgggccgtga tcaccgacga gtacaaggtg 2521 cccagcaaga aattcaaggt gctgggcaac accgaccggc acagcatcaa gaagaacctg 2581 atcggagccc tgctgttcga cagcggcgaa acagccgagg ccacccggct gaagagaacc 2641 gccagaagaa gatacaccag acggaagaac cggatctgct atctgcaaga gatcttcagc 2701 aacgagatgg ccaaggtgga cgacagcttc ttccacagac tggaagagtc cttcctggtg 2761 gaagaggata agaagcacga gcggcacccc atcttcggca acatcgtgga cgaggtggcc 2821 taccacgaga agtaccccac catctaccac ctgagaaaga aactggtgga cagcaccgac 2881 aaggccgacc tgcggctgat ctatctggcc ctggcccaca tgatcaagtt ccggggccac 2941 ttcctgatcg agggcgacct gaaccccgac aacagcgacg tggacaagct gttcatccag 3001 ctggtgcaga cctacaacca gctgttcgag gaaaacccca tcaacgccag cggcgtggac 3061 gccaaggcca tcctgtctgc cagactgagc aagagcagac ggctggaaaa tctgatcgcc 3121 cagctgcccg gcgagaagaa gaatggcctg ttcggaaacc tgattgccct gagcctgggc 3181 ctgaccccca acttcaagag caacttcgac ctggccgagg atgccaaact gcagctgagc 3241 aaggacacct acgacgacga cctggacaac ctgctggccc agatcggcga ccagtacgcc 3301 gacctgtttc tggccgccaa gaacctgtcc gacgccatcc tgctgagcga catcctgaga 3361 gtgaacaccg agatcaccaa ggcccccctg agcgcctcta tgatcaagag atacgacgag 3421 caccaccagg acctgaccct gctgaaagct ctcgtgcggc agcagctgcc tgagaagtac 3481 aaagagattt tcttcgacca gagcaagaac ggctacgccg gctacattga cggcggagcc 3541 agccaggaag agttctacaa gttcatcaag cccatcctgg aaaagatgga cggcaccgag 3601 gaactgctcg tgaagctgaa cagagaggac ctgctgcgga agcagcggac cttcgacaac 3661 ggcagcatcc cccaccagat ccacctggga gagctgcacg ccattctgcg gcggcaggaa 3721 gatttttacc cattcctgaa ggacaaccgg gaaaagatcg agaagatcct gaccttccgc 3781 atcccctact acgtgggccc tctggccagg ggaaacagca gattcgcctg gatgaccaga 3841 aagagcgagg aaaccatcac cccctggaac ttcgaggaag tggtggacaa gggcgcttcc 3901 gcccagagct tcatcgagcg gatgaccaac ttcgataaga acctgcccaa cgagaaggtg 3961 ctgcccaagc acagcctgct gtacgagtac ttcaccgtgt ataacgagct gaccaaagtg 4021 aaatacgtga ccgagggaat gagaaagccc gccttcctga gcggcgagca gaaaaaggcc 4081 atcgtggacc tgctgttcaa gaccaaccgg aaagtgaccg tgaagcagct gaaagaggac 4141 tacttcaaga aaatcgagtg cttcgactcc gtggaaatct ccggcgtgga agatcggttc 4201 aacgcctccc tgggcacata ccacgatctg ctgaaaatta tcaaggacaa ggacttcctg 4261 gacaatgagg aaaacgagga cattctggaa gatatcgtgc tgaccctgac actgtttgag 4321 gacagagaga tgatcgagga acggctgaaa acctatgccc acctgttcga cgacaaagtg 4381 atgaagcagc tgaagcggcg gagatacacc ggctggggca ggctgagccg gaagctgatc 4441 aacggcatcc gggacaagca gtccggcaag acaatcctgg atttcctgaa gtccgacggc 4501 ttcgccaaca gaaacttcat gcagctgatc cacgacgaca gcctgacctt taaagaggac 4561 atccagaaag cccaggtgtc cggccagggc gatagcctgc acgagcacat tgccaatctg 4621 gccggcagcc ccgccattaa gaagggcatc ctgcagacag tgaaggtggt ggacgagctc 4681 gtgaaagtga tgggccggca caagcccgag aacatcgtga tcgaaatggc cagagagaac 4741 cagaccaccc agaagggaca gaagaacagc cgcgagagaa tgaagcggat cgaagagggc 4801 atcaaagagc tgggcagcca gatcctgaaa gaacaccccg tggaaaacac ccagctgcag 4861 aacgagaagc tgtacctgta ctacctgcag aatgggcggg atatgtacgt ggaccaggaa 4921 ctggacatca accggctgtc cgactacgat gtggaccata tcgtgcctca gagctttctg 4981 aaggacgact ccatcgacaa caaggtgctg accagaagcg acaagaaccg gggcaagagc 5041 gacaacgtgc cctccgaaga ggtcgtgaag aagatgaaga actactggcg gcagctgctg 5101 aacgccaagc tgattaccca gagaaagttc gacaatctga ccaaggccga gagaggcggc 5161 ctgagcgaac tggataaggc cggcttcatc aagagacagc tggtggaaac ccggcagatc 5221 acaaagcacg tggcacagat cctggactcc cggatgaaca ctaagtacga cgagaatgac 5281 aagctgatcc gggaagtgaa agtgatcacc ctgaagtcca agctggtgtc cgatttccgg 5341 aaggatttcc agttttacaa agtgcgcgag atcaacaact accaccacgc ccacgacgcc 5401 tacctgaacg ccgtcgtggg aaccgccctg atcaaaaagt accctaagct ggaaagcgag 5461 tttgtgtacg gcgactacaa ggtgtacgac gtgcggaaga tgatcgccaa gagcgagcag 5521 gaaatcggca aggctaccgc caagtacttc ttctacagca acatcatgaa ctttttcaag 5581 accgagatta ccctggccaa cggcgagatc cggaagcggc ctctgatcga gacaaacggc 5641 gaaaccgggg agatcgtgtg ggataagggc cgggattttg ccaccgtgcg gaaagtgctg 5701 agcatgcccc aagtgaatat cgtgaaaaag accgaggtgc agacaggcgg cttcagcaaa 5761 gagtctatcc tgcccaagag gaacagcgat aagctgatcg ccagaaagaa ggactgggac 5821 cctaagaagt acggcggctt cgacagcccc accgtggcct attctgtgct ggtggtggcc 5881 aaagtggaaa agggcaagtc caagaaactg aagagtgtga aagagctgct ggggatcacc 5941 atcatggaaa gaagcagctt cgagaagaat cccatcgact ttctggaagc caagggctac 6001 aaagaagtga aaaaggacct gatcatcaag ctgcctaagt actccctgtt cgagctggaa 6061 aacggccgga agagaatgct ggcctctgcc ggcgaactgc agaagggaaa cgaactggcc 6121 ctgccctcca aatatgtgaa cttcctgtac ctggccagcc actatgagaa gctgaagggc 6181 tcccccgagg ataatgagca gaaacagctg tttgtggaac agcacaagca ctacctggac 6241 gagatcatcg agcagatcag cgagttctcc aagagagtga tcctggccga cgctaatctg 6301 gacaaagtgc tgtccgccta caacaagcac cgggataagc ccatcagaga gcaggccgag 6361 aatatcatcc acctgtttac cctgaccaat ctgggagccc ctgccgcctt caagtacttt 6421 gacaccacca tcgaccggaa gaggtacacc agcaccaaag aggtgctgga cgccaccctg 6481 atccaccaga gcatcaccgg cctgtacgag acacggatcg acctgtctca gctgggaggt 6541 gactctggcg gctcaaaaag aaccgccgac ggcagcgaat tcgagcccaa gaagaagagg 6601 aaagtctgag atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag 6661 catctgactt ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt 6721 ttgtgtctct cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag 6781 tatttggttt agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg 6841 ctataaagag gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga 6901 aaagccttga cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta 6961 acatccctaa aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta 7021 ctcccagtca tagctgtccc tcttctctta tggagatccc tcgacggatc ggccgcaatt 7081 cgtaaccatg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa 7141 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac 7201 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca 7261 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc 7321 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc 7381 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc 7441 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag 7501 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc 7561 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt 7621 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct 7681 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg 7741 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct 7801 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat 7861 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg 7921 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa 7981 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt 8041 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc 8101 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt 8161 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta 8221 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat 8281 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac 8341 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg 8401 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag 8461 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt 8521 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt 8581 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt 8641 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt 8701 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct 8761 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt 8821 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac 8881 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa 8941 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa 9001 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca 9061 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct 9121 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga 9181 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc 9241 taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca 9301 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag 9361 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc 9421 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc 9481 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc 9541 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa 9601 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc 9661 acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcgc 9721 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg 9781 ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt 9841 aaaacgacgg ccagtgagcg cgcgtaatac gactcactat agggcgaatt ggagctccac 9901 cgcggtggcg gccgctctag agagcttggc ccattgcata cgttgtatcc atatcataat 9961 atgtacattt atattggctc atgtccaaca ttaccgccat gttgacattg attattgact 10021 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc 10081 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg 10141 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa 10201 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca 10261 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac 10321 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc 10381 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga 10441 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg 10501 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta 10561 cggtgggagg tctatataag cagagctcgt ttagtgaacc gtcagatcgc ctggagacgc 10621 catccacgct gttttgacct ccatagaaga caccgggacc gatccagcct ccggtcgacc 10681 gatcctgaga acttcagggt gagtttgggg acccttgatt gttctttctt tttcgctatt 10741 gtaaaattca tgttatatgg agggggcaaa gttttcaggg tgttgtttag aatgggaaga 10801 tgtcccttgt atcaccatgg accctcatga taattttgtt tctttcactt tctactctgt 10861 tgacaaccat tgtctcctct tattttcttt tcattttctg taactttttc gttaaacttt 10921 agcttgcatt tgtaacgaat ttttaaattc acttttgttt atttgtcaga ttgtaagtac 10981 tttctctaat cacttttttt tcaaggcaat cagggtatat tatattgtac ttcagcacag 11041 ttttagagaa caattgttat aattaaatga taaggtagaa tatttctgca tataaattct 11101 ggctggcgtg gaaatattct tattggtaga aacaactaca ccctggtcat catcctgcct 11161 ttctctttat ggttacaatg atatacactg tttgagatga ggataaaata ctctgagtcc 11221 aaaccgggcc cctctgctaa ccatgttcat gccttcttct ctttcctaca gctcctgggc 11281 aacgtgctgg ttgttgtgct gtctcatcat tttggcaaag aattcctcga gggccggatc 11341 cgaattcctg tatttgtctg aaaac //