pWS172

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联系方式:I47-825O-882O

相关技术服务:质粒构建    基因合成    质粒大提

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载体基本信息

质粒名称: pWS172
出品公司: Addgene
目录编号: 90519
存储实验室: Tom Ellis

载体骨架基本信息
载体名称: 2 micron
空载体大小: 3882 bp
完整质粒大小: 11132bp
载体类型: Yeast Expression, CRISPR
筛选标记: HIS3
载体抗性: Kanamycin
生长温度: 37 ℃
宿主菌: NEB Stable
拷贝数: 高拷贝

插入基因信息
插入基因名称: Cas9
物种来源: S. cerevisiae (budding yeast)

文献引用方法
材料和方法部分: pWS172 was a gift from Tom Ellis (Addgene plasmid # 90519 ; http://n2t.net/addgene:90519 ; RRID:Addgene_90519)

载体图谱质粒图谱和多克隆位点信息

pWS172载体图谱

载体简介

载体序列

LOCUS       pWS172               11132 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Cas9 gap repair vector - HIS3.
ACCESSION   .
VERSION     .
KEYWORDS    pWS172
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11132)
  TITLE     Ellis Lab Yeast CRISPR Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 11132)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.biofeng.com
FEATURES             Location/Qualifiers
     source          1..11132
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             1451..5554
                     /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             5567..5587
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5591..5611
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     RBS             6521..6532
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             6539..7252
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      7264..7335
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      7351..7378
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_RNA        7392..7467
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        7988..8175
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             8176..8838
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      9053..10035
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     CDS             complement(10093..10908)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     10816..10835
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     10900..10919
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtgc ggccgcccct gaattcgcat ctagactgaa
      721 ctggccgata attgcagacg aacggtgagt aaggaaagag tgaggaacta tcgcatacct
      781 gcatttaaag atgccgattt gggcgcgaat cctttatttt ggcttcaccc tcatactatt
      841 atcagggcca gaaaaaggaa gtgtttccct ccttcttgaa ttgatgttac cctcataaag
      901 cacgtggcct cttatcgaga aagaaattac cgtcgctcgt gatttgtttg caaaaagaac
      961 aaaactgaaa aaacccagac acgctcgact tcctgtcatc ctattgattg cagcttccaa
     1021 tttcgtcaca caacaaggtc ctagcgacgg ctcacaggtt ttgtaacaag caatcgaagg
     1081 ttctggaatg gcgggaaagg gtttagtacc acatgctatg atgcccactg tgatctccag
     1141 agcaaagttc gttcgatcgt actgttactc tctctctttc aaacagaatt gtccgaatcg
     1201 tgtgacaaca acagcctgtt ctcacacact cttttcttct aaccaagggg gtggtttagt
     1261 ttagtagaac ctcgtgaaac ttacatttac atatatataa acttgcataa attggtcaat
     1321 gcaagaaata catatttggt cttttctaat tcgtagtttt tcaagttctt agatgctttc
     1381 tttttctctt ttttacagat catcaaggaa gtaattatct actttttaca acaaatataa
     1441 aacaagatct atggacaaga agtattctat cggactggat atcgggacta atagcgtcgg
     1501 gtgggccgtg atcactgacg agtacaaggt gccctctaag aagttcaagg tgctcgggaa
     1561 caccgaccgg cattccatca agaaaaatct gatcggagct ctcctctttg attcagggga
     1621 aaccgctgaa gcaacccgcc tcaagcggac tgctagacgg cggtacacca ggaggaagaa
     1681 ccggatttgt taccttcaag agatattctc caacgaaatg gcaaaggtcg acgacagctt
     1741 cttccatagg ctggaagaat cattcctcgt ggaagaggat aagaagcatg aacggcatcc
     1801 catcttcggt aatatcgtcg acgaggtggc ctatcacgag aaatacccaa ccatctacca
     1861 tcttcgcaaa aagctggtgg actcaaccga caaggcagac ctccggctta tctacctggc
     1921 cctggcccac atgatcaagt tcagaggcca cttcctgatc gagggcgacc tcaatcctga
     1981 caatagcgat gtggataaac tgttcatcca gctggtgcag acttacaacc agctctttga
     2041 agagaacccc atcaatgcaa gcggagtcga tgccaaggcc attctgtcag cccggctgtc
     2101 aaagagccgc agacttgaga atcttatcgc tcagctgccg ggtgaaaaga aaaatggact
     2161 gttcgggaac ctgattgctc tttcacttgg gctgactccc aatttcaagt ctaatttcga
     2221 cctggcagag gatgccaagc tgcaactgtc caaggacacc tatgatgacg atctcgacaa
     2281 cctcctggcc cagatcggtg accaatacgc cgaccttttc cttgctgcta agaatctttc
     2341 tgacgccatc ctgctgtctg acattctccg cgtgaacact gaaatcacca aggcccctct
     2401 ttcagcttca atgattaagc ggtatgatga gcaccaccag gacctgaccc tgcttaaggc
     2461 actcgtccgg cagcagcttc cggagaagta caaggaaatc ttctttgacc agtcaaagaa
     2521 tggatacgcc ggctacatcg acggaggtgc ctcccaagag gaattttata agtttatcaa
     2581 acctatcctt gagaagatgg acggcaccga agagctcctc gtgaaactga atcgggagga
     2641 tctgctgcgg aagcagcgca ctttcgacaa tgggagcatt ccccaccaga tccatcttgg
     2701 ggagcttcac gccatccttc ggcgccaaga ggacttctac ccctttctta aggacaacag
     2761 ggagaagatt gagaaaattc tcactttccg catcccctac tacgtgggac ccctcgccag
     2821 aggaaatagc cggtttgctt ggatgaccag aaagtcagaa gaaactatca ctccctggaa
     2881 cttcgaagag gtggtggaca agggagccag cgctcagtca ttcatcgaac ggatgactaa
     2941 cttcgataag aacctcccca atgagaaggt cctgccgaaa cattccctgc tctacgagta
     3001 ctttaccgtg tacaacgagc tgaccaaggt gaaatatgtc accgaaggga tgaggaagcc
     3061 cgcattcctg tcaggcgaac aaaagaaggc aattgtggac cttctgttca agaccaatag
     3121 aaaggtgacc gtgaagcagc tgaaggagga ctatttcaag aaaattgaat gcttcgactc
     3181 tgtggagatt agcggggtcg aagatcggtt caacgcaagc ctgggtactt accatgatct
     3241 gcttaagatc atcaaggaca aggattttct ggacaatgag gagaacgagg acatccttga
     3301 ggacattgtc ctgactctca ctctgttcga ggaccgggaa atgatcgagg agaggcttaa
     3361 gacctacgcc catctgttcg acgataaagt gatgaagcaa cttaaacgga gaagatatac
     3421 cggatgggga cgccttagcc gcaaactcat caacggaatc cgggacaaac agagcggaaa
     3481 gaccattctt gatttcctta agagcgacgg attcgctaat cgcaacttca tgcaacttat
     3541 ccatgatgat tccctgacct ttaaggagga catccagaag gcccaagtgt ctggacaagg
     3601 tgactcactg cacgagcata tcgcaaatct ggctggttca cccgctatta agaagggtat
     3661 tctccagacc gtgaaagtcg tggacgagct ggtcaaggtg atgggtcgcc ataaaccaga
     3721 gaacattgtc atcgagatgg ccagggaaaa ccagactacc cagaagggac agaagaacag
     3781 cagggagcgg atgaaaagaa ttgaggaagg gattaaggag ctcgggtcac agatccttaa
     3841 agagcacccg gtggaaaaca cccagcttca gaatgagaag ctctatctgt actaccttca
     3901 aaatggacgc gatatgtatg tggaccaaga gcttgatatc aacaggctct cagactacga
     3961 cgtggaccat atcgtccctc agagcttcct caaagacgac tcaattgaca ataaggtgct
     4021 gactcgctca gacaagaacc ggggaaagtc agataacgtg ccctcagagg aagtcgtgaa
     4081 aaagatgaag aactattggc gccagcttct gaacgcaaag ctgatcactc agcggaagtt
     4141 cgacaatctc actaaggctg agaggggcgg actgagcgaa ctggacaaag caggattcat
     4201 taaacggcaa cttgtggaga ctcggcagat tactaaacat gtcgcccaaa tccttgactc
     4261 acgcatgaat accaagtacg acgaaaacga caaacttatc cgcgaggtga aggtgattac
     4321 cctgaagtcc aagctggtca gcgatttcag aaaggacttt caattctaca aagtgcggga
     4381 gatcaataac tatcatcatg ctcatgacgc atatctgaat gccgtggtgg gaaccgccct
     4441 gatcaagaag tacccaaagc tggaaagcga gttcgtgtac ggagactaca aggtctacga
     4501 cgtgcgcaag atgattgcca aatctgagca ggagatcgga aaggccaccg caaagtactt
     4561 cttctacagc aacatcatga atttcttcaa gaccgaaatc acccttgcaa acggtgagat
     4621 ccggaagagg ccgctcatcg agactaatgg ggagactggc gaaatcgtgt gggacaaggg
     4681 cagagatttc gctaccgtgc gcaaagtgct ttctatgcct caagtgaaca tcgtgaagaa
     4741 aaccgaggtg caaaccggag gcttttctaa ggaatcaatc ctccccaagc gcaactccga
     4801 caagctcatt gcaaggaaga aggattggga ccctaagaag tacggcggat tcgattcacc
     4861 aactgtggct tattctgtcc tggtcgtggc taaggtggaa aaaggaaagt ctaagaagct
     4921 caagagcgtg aaggaactgc tgggtatcac cattatggag cgcagctcct tcgagaagaa
     4981 cccaattgac tttctcgaag ccaaaggtta caaggaagtc aagaaggacc ttatcatcaa
     5041 gctcccaaag tatagcctgt tcgaactgga gaatgggcgg aagcggatgc tcgcctccgc
     5101 tggcgaactt cagaagggta atgagctggc tctcccctcc aagtacgtga atttcctcta
     5161 ccttgcaagc cattacgaga agctgaaggg gagccccgag gacaacgagc aaaagcaact
     5221 gtttgtggag cagcataagc attatctgga cgagatcatt gagcagattt ccgagttttc
     5281 taaacgcgtc attctcgctg atgccaacct cgataaagtc cttagcgcat acaataagca
     5341 cagagacaaa ccaattcggg agcaggctga gaatatcatc cacctgttca ccctcaccaa
     5401 tcttggtgcc cctgccgcat tcaagtactt cgacaccacc atcgaccgga aacgctatac
     5461 ctccaccaaa gaagtgctgg acgccaccct catccaccag agcatcaccg gactttacga
     5521 aactcggatt gacctctcac agctcggagg ggatgaggga gctgatccaa aaaagaagag
     5581 aaaggtagat ccaaaaaaga agagaaaggt ataaggatcc taactcgaga ttgaattgaa
     5641 ttgaaatcga tagatcaatt tttttctttt ctctttcccc atcctttacg ctaaaataat
     5701 agtttatttt attttttgaa tattttttat ttatatacgt atatatagac tattatttat
     5761 cttttaatga ttattaagat ttttattaaa aaaaaattcg ctcctctttt aatgccttta
     5821 tgcagttttt ttttcccatt cgatatttct atgtgctgcc aagatatcaa caacagagga
     5881 catatgccct acctccatga tctactgacg tccctgaggc tgcaattcat gtaatggggc
     5941 agtatccgcg gcaagtccta gtgcaatggc ggttttttac cctcgttctg aagaagaggc
     6001 gacgcgggtg cggtcatcac taatgtggaa attggtaaga ctctcgggcc tccgccttta
     6061 ggcggtgctt actctttcat aaaggggctg ttagttatgc aacgatgtgc ttcagtatta
     6121 cattttttgc cttcaacgcc ttgattgttc tatttttgct aataataaat ctatttcatc
     6181 ggactaaaag tccattagtt gtaagcggat ttagctcagt tgggagagcg ccagactgaa
     6241 gaaaaacttc ggtcaagtca tctggaggtc ctgtgttcga tccacagaat tcgcagatgg
     6301 ccggcatggt cccagcctcc tcgctggcgc cggctgggca acaccttcgg gtggcgaatg
     6361 ggacttgaga cggaaagtga aacgtgattt catgcgtcat tttgaacatt ttgtaaatct
     6421 tatttaataa tgtgtgcggc aattcacatt taatttatga atgttttctt aacatcgcgg
     6481 caactcaaga aacggcaggt tcggatctta gctactagag aaagaggaga aatactagat
     6541 gcgtaaaggc gaagagctgt tcactggtgt cgtccctatt ctggtggaac tggatggtga
     6601 tgtcaacggt cataagtttt ccgtgcgtgg cgagggtgaa ggtgacgcaa ctaatggtaa
     6661 actgacgctg aagttcatct gtactactgg taaactgccg gttccttggc cgactctggt
     6721 aacgacgctg acttatggtg ttcagtgctt tgctcgttat ccggaccata tgaagcagca
     6781 tgacttcttc aagtccgcca tgccggaagg ctatgtgcag gaacgcacga tttcctttaa
     6841 ggatgacggc acgtacaaaa cgcgtgcgga agtgaaattt gaaggcgata ccctggtaaa
     6901 ccgcattgag ctgaaaggca ttgactttaa agaggacggc aatatcctgg gccataagct
     6961 ggaatacaat tttaacagcc acaatgttta catcaccgcc gataaacaaa aaaatggcat
     7021 taaagcgaat tttaaaattc gccacaacgt ggaggatggc agcgtgcagc tggctgatca
     7081 ctaccagcaa aacactccaa tcggtgatgg tcctgttctg ctgccagaca atcactatct
     7141 gagcacgcaa agcgttctgt ctaaagatcc gaacgagaaa cgcgatcata tggttctgct
     7201 ggagttcgta accgcagcgg gcatcacgca tggtatggat gaactgtaca aatgaccagg
     7261 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     7321 tcggtgaacg ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt
     7381 tatacgtctc agttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa
     7441 cttgaaaaag tggcaccgag tcggtgcttt ttttattttt tgtcactatt gttatgtaaa
     7501 atgccacctc tgacagtatg gaacgcaaac ttctgtctag tggatagctg gacattccag
     7561 gcagtgcctc tgccgccgga cccctctcgt gattgggtag ctggacatgc ccttgtaaga
     7621 tataacaaga gcctgcctgt ctaatgatct cacggcgaaa gtcggggaga cagcagcggc
     7681 tgcagacatt ataccgcaac aacactaagg tgagataact ccgtacattg actacgcgtt
     7741 cctctagacc ttacttgacc ggatacagtg tctttgacac gtttatgggt tacagcaatc
     7801 acatccaaga ctggctatgc acgaagcaac tcttgagtgt taaaatgttg acccctgtat
     7861 ttgggatgcg ggtagtagat gagtgcagga tatcagcagt tacagagatg ttacgaacca
     7921 ctagtgcact gcagtacaaa cacagtcctt tcccgcaatt ttctttttct attactcttg
     7981 gcctcctcta gtacactcta tattttttta tgcctcggta atgattttca tttttttttt
     8041 tccacctagc ggatgactct ttttttttct tagcgattgg cattatcaca taatgaatta
     8101 tacattatat aaagtaatgt gatttcttcg aagaatatac taaaaaatga gcaggcaaga
     8161 taaacgaagg caaagatgac agagcagaaa gccctagtaa agcgtattac aaatgaaacc
     8221 aagattcaga ttgcgatctc tttaaagggt ggtcccctag cgatagagca ctcgatcttc
     8281 ccagaaaaag aggcagaagc agtagcagaa caggccacac aatcgcaagt gattaacgtc
     8341 cacacaggta tagggtttct ggaccatatg atacatgctc tggccaagca ttccggctgg
     8401 tcgctaatcg ttgagtgcat tggtgactta cacatagacg accatcacac cactgaggac
     8461 tgcgggattg ctctcggtca agcttttaaa gaggccctag gggccgtgcg tggagtaaaa
     8521 aggtttggat caggatttgc gcctttggat gaggcacttt ccagagcggt ggttgatctt
     8581 tcgaacaggc cgtacgcagt tgtcgaactt ggtttgcaaa gggagaaagt aggtgatctc
     8641 tcttgcgaga tgatcccgca ttttcttgaa agctttgcag aggctagcag aattaccctc
     8701 cacgttgatt gtctgcgagg caagaatgat catcaccgta gtgagagtgc gttcaaggct
     8761 cttgcggttg ccataagaga agccacctcg cccaatggta ccaacgatgt tccctccacc
     8821 aaaggtgttc ttatgtagtg acaccgatta tttaaagctg ctgcatacga tatatataca
     8881 tgtgtatata tgtataccta tgaatgtcag taagtatgta tacgaacagt atgatactga
     8941 agatgacaag gtaatgcatc attctatacg tgtcattctg aacgaggcgc gctttccttt
     9001 tttctttttg ctttttcttt ttttttctct tgaactcgac ggatcataga gtaacgaagc
     9061 atctgtgctt cattttgtag aacaaaaatg caacgcgaga gcgctaattt ttcaaacaaa
     9121 gaatctgagc tgcattttta cagaacagaa atgcaacgcg aaagcgctat tttaccaacg
     9181 aagaatctgt gcttcatttt tgtaaaacaa aaatgcaacg cgagagcgct aatttttcaa
     9241 acaaagaatc tgagctgcat ttttacagaa cagaaatgca acgcgagagc gctattttac
     9301 caacaaagaa tctatacttc ttttttgttc tacaaaaatg catcccgaga gcgctatttt
     9361 tctaacaaag catcttagat tacttttttt ctcctttgtg cgctctataa tgcagtctct
     9421 tgataacttt ttgcactgta ggtccgttaa ggttagaaga aggctacttt ggtgtctatt
     9481 ttctcttcca taaaaaaagc ctgactccac ttcccgcgtt tactgattac tagcgaagct
     9541 gcgggtgcat tttttcaaga taaaggcatc cccgattata ttctataccg atgtggattg
     9601 cgcatacttt gtgaacagaa agtgatagcg ttgatgattc ttcattggtc agaaaattat
     9661 gaacggtttc ttctattttg tctctatata ctacgtatag gaaatgttta cattttcgta
     9721 ttgttttcga ttcactctat gaatagttct tactacaatt tttttgtcta aagagtaata
     9781 ctagagataa acataaaaaa tgtagaggtc gagtttagat gcaagttcaa ggagcgaaag
     9841 gtggatgggt aggttatata gggatatagc acagagatat atagcaaaga gatacttttg
     9901 agcaatgttt gtggaagcgg tattcgcaat attttagtag ctcgttacag tccggtgcgt
     9961 ttttggtttt ttgaaagtgc gtcatcagag cgcttttggt tttcaaaagc gctctgaagt
    10021 tcctatactt tctagctaga gaataggaac ttcccgagcg gccgcgtgtt acaaccaatt
    10081 aaccaattct gattagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc
    10141 aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc
    10201 gaggcagttc cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac
    10261 atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc
    10321 atgagtgacg actgaatccg gtgagaatgg caaaagctta tgcatttctt tccagacttg
    10381 ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt
    10441 cattcgtgat tgcgcctgag cgaggcgaaa tacgcgatcg ctgttaaaag gacaattaca
    10501 aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc
    10561 tgaatcagga tattcttcta atacctggaa tgctgttttc ccggggatcg cagtggtgag
    10621 taaccatgca tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc
    10681 cgtcagccag tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc
    10741 atgtttcaga aacaactctg gcgcatcggg cttcccatac aatcgataga ttgtcgcacc
    10801 tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga
    10861 atttaatcgc ggcctggagc aagacgtttc ccgttgaata tggctcataa caccccttgt
    10921 attactgttt atgtaagcag acagttttat tgttcatgat gatatatttt tatcttgtgc
    10981 aatgtaacat cagagatttt gagacacaac gtggctttgt tgaataaatc gaacttttgc
    11041 tgagttgaag gatcagtcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
    11101 tcagaccccg tagaaaagat caaaggatct tc
//

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