pSLQ9926: pHR-PGK-SV40_NLS-dCasMINI-V4-VPR-c- Myc_NLS-mCherr

价格:1200元

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: pSLQ9926: pHR-PGK-SV40_NLS-dCasMINI-V4-VPR-c- Myc_NLS-mCherry-WPRE
出品公司: Addgene
目录编号: 176269
存储实验室: Stanley Qi

载体骨架基本信息
载体名称: pHR
载体出品公司: --
空载体大小: --
完整质粒大小: --
载体修饰: --
载体类型: 哺乳动物表达,慢病毒
筛选标记: --
载体抗性: 氨苄霉素
生长温度: 37 ℃
宿主菌: NEB Stable
拷贝数: --

插入基因信息
插入基因名称: dCasMINI-V4-VPR-mCherry
物种来源: --
插入大小: --
基因突变: --
GenBank编号 --
启动子: PGK
标签/融合蛋白: --
克隆方法: Gibson克隆
5’克隆位点: --
3’克隆位点: --
5’测序引物 CAAAAGCGCACGTCTGCCGC
3’测序引物 --

文献引用方法
材料和方法部分: pSLQ9926: pHR-PGK-SV40_NLS-dCasMINI-V4-VPR-c- Myc_NLS-mCherry-WPRE was a gift from Stanley Qi (Addgene plasmid # 176269 ; http://n2t.net/addgene:176269 ; RRID:Addgene_176269)
参考文献部分: Engineered miniature CRISPR-Cas system for mammalian genome regulation and editing. Xu X, Chemparathy A, Zeng L, Kempton HR, Shang S, Nakamura M, Qi LS. Mol Cell. 2021 Aug 26. pii: S1097-2765(21)00648-1. doi: 10.1016/j.molcel.2021.08.008. 10.1016/j.molcel.2021.08.008 PubMed 34480847

序号

载体图谱质粒图谱和多克隆位点信息

pSLQ9926pHR-PGK-SV40_NLS-dCasMINI-V4-VPR-c- Myc_NLS-mCherry-WPRE质粒图谱

载体简介

载体序列

LOCUS       pSLQ9926: pHR-PGK-SV40_NLS-dCasMINI-V4-VPR-c- Myc_NLS-mCherry-...              13028 bp ds-DNA     circular SYN 01-OCT-2021
DEFINITION  Expression of dCasMINI-V4-VPR-mCherry in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pSLQ9926: pHR-PGK-SV40_NLS-dCasMINI-V4-VPR-c- Myc_NLS-mCherry-...
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13028)
  AUTHORS   Xu X, Chemparathy A, Zeng L, Kempton HR, Shang S, Nakamura M, Qi LS
  TITLE     Engineered miniature CRISPR-Cas system for mammalian genome 
            regulation and editing.
  JOURNAL   Mol Cell. 2021 Aug 26. pii: S1097-2765(21)00648-1. doi: 
            10.1016/j.molcel.2021.08.008.
  PUBMED    34480847
REFERENCE   2  (bases 1 to 13028)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 1, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..13028
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    27..144
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        209..708
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(244..267)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     641..660
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      768..777
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             780..800
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2400..2426
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2478..2627
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             3072..3443
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALLSQISS"
     CDS             3462..4031
                     /codon_start=1
                     /product="transcriptional activation domain from the human 
                     herpesvirus 4 (Epstein-Barr virus) replication and 
                     transcription activator Rta/BRLF1 (Hardwick et al., 1992; 
                     Chavez et al., 2015)"
                     /label=Rta AD
                     /translation="RDSREGMFLPKPEAGSAISDVFEGREVCQPKRIRPFHPPGSPWAN
                     RPLPASLAPTPTGPVHEPVGSLTPAPVPQPLDPAPAVTPEASHLLEDPDEETSQAVKAL
                     REMADTVIPQKEEAAICGQMDLSHPPPRGHLDELTTTLESMTEDLNLDSPLTPELNEIL
                     DTFLNDECLLHAMHISTGLSIFDTSLF"
     CDS             4059..4085
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             4098..4805
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(4240..4258)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4508..4527
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    4869..5457
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4922..4942)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5340..5351)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5460..5476)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5461..5477)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5567..5800
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     promoter        complement(5825..5843)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5826..5843)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(6133..6152)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(6366..6385)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6485..6507
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6545..6563)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6631..6735
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6736..7596
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6954..6973)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7767..8355
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8256..8275
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8509..8526
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(8596..8616)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        8601..8930
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      8781..8916
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     8843..8862
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     intron          10064..10129
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             10259..10279
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    10704..10838
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10741..10760)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10795..10814
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             11006..11639
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    11686..11811
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    12308..12541
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             12726..12770
                     /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 cgccaaatgg cagtattcat ccacaatttt aaaagaaaag gggggattgg ggggtacagt
       61 gcaggggaaa gaatagtaga cataatagca acagacatac aaactaaaga attacaaaaa
      121 caaattacaa aaattcaaaa ttttcgggtt tattacaggg acagcagaga tccagtttgg
      181 atcgataagc ttgatatcga attctaccgg gtaggggagg cgcttttccc aaggcagtct
      241 ggagcatgcg ctttagcagc cccgctgggc acttggcgct acacaagtgg cctctggcct
      301 cgcacacatt ccacatccac cggtaggcgc caaccggctc cgttctttgg tggccccttc
      361 gcgccacctt ctactcctcc cctagtcagg aagttccccc ccgccccgca gctcgcgtcg
      421 tgcaggacgt gacaaatgga agtagcacgt ctcactagtc tcgtgcagat ggacagcacc
      481 gctgagcaat ggaagcgggt aggcctttgg ggcagcggcc aatagcagct ttgctccttc
      541 gctttctggg ctcagaggct gggaaggggt gggtccgggg gcgggctcag gggcgggctc
      601 aggggcgggg cgggcgcccg aaggtcctcc ggaggcccgg cattctgcac gcttcaaaag
      661 cgcacgtctg ccgcgctgtt ctcctcttcc tcatctccgg gcctttcgac ctgcagccca
      721 agcttacgcg tctgcaggat atcaagcttg cggtaccgcg ggcccgggcc accatgggac
      781 ccaagaaaaa acgcaaggtg ggaagcggat ccgccaaaaa caccattacc aaaacactga
      841 aactgcgtat tgtgcgtccg tataatagcg cagaagtgga aaaaattgtt gccgacgaaa
      901 aaaacaaccg cgaaaaaatc gcactggaaa agaacaaaga caaagtgaaa gaagcctgca
      961 gcaaacatct gaaagttgca gcatattgta ccacacaggt tgaacgtaat gcatgcctgt
     1021 tttgtaaagc acgtaaactg gatgacaaat tctaccaaaa actgcgtggt cagtttccgg
     1081 atgcagtttt ttggcaagaa atcagcgaaa tttttcgcca gctgcagaaa caggcagcag
     1141 aaatctataa tcagagcctg atcgaactgt actacgagat ttttatcaaa ggcaaaggta
     1201 ttgcaaatgc cagcagcgtt gaacattatc tgagtagagt ttgttataga cgtgcagcag
     1261 aactgtttaa aaacgcagca attgcaagcg gtctgcgtag caaaatcaaa agcaattttc
     1321 gtctgaaaga actgaaaaac atgaaaagtg gtctgccgac caccaaaagc gataattttc
     1381 cgattccgct ggttaaacag aaaggtggtc agtataccgg ttttgaaatt agcaatcata
     1441 atagcgactt catcatcaag attccgtttg gtcgttggca ggtcaaaaaa gagattgata
     1501 aatatcgtcc gtgggagaaa tttgactttg aacaggttca gaaaagcccg aaaccgatta
     1561 gcctgctgct gagcacccag cgtcgtaaac gtaataaagg ttggagcaaa gatgaaggca
     1621 ccgaagccga aatcaaaaaa gttatgaatg gcgattatca gaccagctac attgaagtta
     1681 aacgtggcag caaaatctgt gaaaaaagcg catggatgct gaatctgagc attgatgttc
     1741 cgaaaattga taaaggtgtg gatccgagca ttattggtgg tattgcagtt ggtgttagat
     1801 caccgctggt ttgcgcaatt aacaatgcat ttagccgtta tagcatcagc gataacgacc
     1861 tgtttcactt caacaagaaa atgtttgcac gtcgtcgtat cctgctgaaa aaaaaccgtc
     1921 ataaacgtgc aggtcatggt gcaaaaaaca aactgaaacc gatcaccatt ctgaccgaaa
     1981 aaagtgaacg ttttcgcaaa aagctgattg aacgttgggc atgtgaaatc gcggatttct
     2041 tcattaaaaa caaagttggc accgtgcaga tggaaaatct ggaaagcatg aaacgtaaag
     2101 aggacagcta ttttaacatt cgcctgcgtg gcttttggcc gtatgcagaa atgcagaaca
     2161 aaatcgaatt caaactgaag cagtatggca tcgaaattcg taaagttgca ccgaataata
     2221 ccagcaaaac ctgtagcaaa tgtggccatc tgaacaacta tttcaacttc gagtaccgca
     2281 agaaaaacaa attcccgcac tttaaatgcg aaaaatgcaa cttcaaagaa aacgccgcgt
     2341 ataatgcagc cctgaatatt tcaaacccga aactgaaaag caccaaagag agaccggcct
     2401 atccctatga cgtgcccgat tatgccagcc tgggcagcgg cgacggcatt ggtagtggga
     2461 gcaacggcag cagcctcgat gctttagacg attttgactt agatatgctt ggttcagacg
     2521 cgttagacga cttcgaccta gacatgttag gctcagatgc attggacgac ttcgatttag
     2581 atatgttggg ctccgatgcc ctagatgact ttgatctaga tatgctaggt agtggcggca
     2641 gcggatccca gtatctgccc gacacagatg atagacaccg aatcgaagag aaacgcaagc
     2701 gaacgtatga aaccttcaaa tcgatcatga agaaatcgcc cttctcgggt ccgaccgatc
     2761 ccaggccccc accgagaagg attgcggtcc cgtcccgctc gtcggccagc gtgccgaagc
     2821 ctgcgccgca gccctacccc ttcacgtcga gcctgagcac aatcaattat gacgagttcc
     2881 cgacgatggt gttcccctcg ggacaaatct cacaagcctc ggcgctcgca ccagcgcctc
     2941 cccaagtcct tccgcaagcg cctgccccag cgcctgcacc ggcaatggtg tccgccctcg
     3001 cacaggcccc tgcgcccgtc cccgtgctcg cgcctggacc gccccaggcg gtcgctccac
     3061 cggctccgaa gccgacgcag gccggagagg gaacactctc cgaagcactt cttcaactcc
     3121 agtttgatga cgaggatctt ggagcactcc ttggaaactc gacagaccct gcggtgttta
     3181 ccgacctcgc gtcagtagat aactccgaat ttcagcagct tttgaaccag ggtatcccgg
     3241 tcgcgccaca tacaacggag cccatgttga tggaataccc cgaagcaatc acgagacttg
     3301 tgacgggagc gcagcggcct cccgatcccg cacccgcacc tttgggggca cctggcctcc
     3361 ctaacggact tttgagcggc gacgaggatt tctcctccat cgccgatatg gatttctcag
     3421 ccttgctgtc acagatttcc agcggctctg gcagcggcag ccgggattcc agggaaggga
     3481 tgtttttgcc gaagcctgag gccggctccg ctattagtga cgtgtttgag ggccgcgagg
     3541 tgtgccagcc aaaacgaatc cggccatttc atcctccagg aagtccatgg gccaaccgcc
     3601 cactccccgc cagcctcgca ccaacaccaa ccggtccagt acatgagcca gtcgggtcac
     3661 tgaccccggc accagtccct cagccactgg atccagcgcc cgcagtgact cccgaggcca
     3721 gtcacctgtt ggaggatccc gatgaagaga cgagccaggc tgtcaaagcc cttcgggaga
     3781 tggccgatac tgtgattccc cagaaggaag aggctgcaat ctgtggccaa atggaccttt
     3841 cccatccgcc cccaaggggc catctggatg agctgacaac cacacttgag tccatgaccg
     3901 aggatctgaa cctggactca cccctgaccc cggaattgaa cgagattctg gataccttcc
     3961 tgaacgacga gtgcctcttg catgccatgc atatcagcac aggactgtcc atcttcgaca
     4021 catctctgtt tcatatgggt ggaggctccg gggaagatcc tgctgccaaa cgcgttaaac
     4081 tagacatggg aagcggagtg agcaagggcg aggaggataa catggccatc atcaaggagt
     4141 tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg
     4201 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     4261 gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg
     4321 cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct
     4381 tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact
     4441 cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct
     4501 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt
     4561 accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg
     4621 gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg
     4681 gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg
     4741 tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca
     4801 agtaagcggc cgcataagac tctagagtcg acctgcaggc atgcaagctt gatatcaagc
     4861 ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     4921 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     4981 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     5041 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     5101 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     5161 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     5221 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
     5281 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     5341 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     5401 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
     5461 gataccgtcg acctcgaggg aattaattcg agctcggtac ctttaagacc aatgacttac
     5521 aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt
     5581 cactcccaac gaagacaaga tctgcttttt gcttgtactg ggtctctctg gttagaccag
     5641 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
     5701 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
     5761 tccctcagac ccttttagtc agtgtggaaa atctctagca gcatctagaa ttaattccgt
     5821 gtattctata gtgtcaccta aatcgtatgt gtatgataca taaggttatg tattaattgt
     5881 agccgcgttc taacgacaat atgtacaagc ctaattgtgt agcatctggc ttactgaagc
     5941 agaccctatc atctctctcg taaactgccg tcagagtcgg tttggttgga cgaaccttct
     6001 gagtttctgg taacgccgtc ccgcacccgg aaatggtcag cgaaccaatc agcagggtca
     6061 tcgctagcca gatcctctac gccggacgca tcgtggccgg catcaccggc gccacaggtg
     6121 cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct cgccacttcg
     6181 ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc gggggactgt
     6241 tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc
     6301 tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga atggtgcact
     6361 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     6421 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     6481 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     6541 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     6601 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     6661 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     6721 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     6781 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     6841 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     6901 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     6961 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     7021 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     7081 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     7141 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     7201 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     7261 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     7321 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     7381 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     7441 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     7501 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     7561 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     7621 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     7681 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     7741 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     7801 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     7861 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     7921 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     7981 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     8041 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     8101 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     8161 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     8221 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     8281 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     8341 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     8401 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     8461 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     8521 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     8581 cattaatgca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     8641 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     8701 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     8761 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     8821 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     8881 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagcttggac
     8941 acaagacagg cttgcgagat atgtttgaga ataccacttt atcccgcgtc agggagaggc
     9001 agtgcgtaaa aagacgcgga ctcatgtgaa atactggttt ttagtgcgcc agatctctat
     9061 aatctcgcgc aacctatttt cccctcgaac actttttaag ccgtagataa acaggctggg
     9121 acacttcaca tgagcgaaaa atacatcgtc acctgggaca tgttgcagat ccatgcacgt
     9181 aaactcgcaa gccgactgat gccttctgaa caatggaaag gcattattgc cgtaagccgt
     9241 ggcggtctgt accgggtgcg ttactggcgc gtgaactggg tattcgtcat gtcgataccg
     9301 tttgtatttc cagctacgat cacgacaacc agcgcgagct taaagtgctg aaacgcgcag
     9361 aaggcgatgg cgaaggcttc atcgttattg atgacctggt ggataccggt ggtactgcgg
     9421 ttgcgattcg tgaaatgtat ccaaaagcgc actttgtcac catcttcgca aaaccggctg
     9481 gtcgtccgct ggttgatgac tatgttgttg atatcccgca agatacctgg attgaacagc
     9541 cgtgggatat gggcgtcgta ttcgtcccgc caatctccgg tcgctaatct tttcaacgcc
     9601 tggcactgcc gggcgttgtt ctttttaact tcaggcgggt tacaatagtt tccagtaagt
     9661 attctggagg ctgcatccat gacacaggca aacctgagcg aaaccctgtt caaaccccgc
     9721 tttaaacatc ctgaaacctc gacgctagtc cgccgcttta atcacggcgc acaaccgcct
     9781 gtgcagtcgg cccttgatgg taaaaccatc cctcactggt atcgcatgat taaccgtctg
     9841 atgtggatct ggcgcggcat tgacccacgc gaaatcctcg acgtccaggc acgtattgtg
     9901 atgagcgatg ccgaacgtac cgacgatgat ttatacgata cggtgattgg ctaccgtggc
     9961 ggcaactgga tttatgagtg ggccccggat ctttgtgaag gaaccttact tctgtggtgt
    10021 gacataattg gacaaactac ctacagagat ttaaagctct aaggtaaata taaaattttt
    10081 aagtgtataa tgtgttaaac tactgattct aattgtttgt gtattttaga ttccaaccta
    10141 tggaactgat gaatgggagc agtggtggaa tgcctttaat gaggaaaacc tgttttgctc
    10201 agaagaaatg ccatctagtg atgatgaggc tactgctgac tctcaacatt ctactcctcc
    10261 aaaaaagaag agaaaggtag aagaccccaa ggactttcct tcagaattgc taagtttttt
    10321 gagtcatgct gtgtttagta atagaactct tgcttgcttt gctatttaca ccacaaagga
    10381 aaaagctgca ctgctataca agaaaattat ggaaaaatat tctgtaacct ttataagtag
    10441 gcataacagt tataatcata acatactgtt ttttcttact ccacacaggc atagagtgtc
    10501 tgctattaat aactatgctc aaaaattgtg tacctttagc tttttaattt gtaaaggggt
    10561 taataaggaa tatttgatgt atagtgcctt gactagagat cataatcagc cataccacat
    10621 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
    10681 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
    10741 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
    10801 tgtccaaact catcaatgta tcttatcatg tctggatcaa ctggataact caagctaacc
    10861 aaaatcatcc caaacttccc accccatacc ctattaccac tgccaattac ctgtggtttc
    10921 atttactcta aacctgtgat tcctctgaat tattttcatt ttaaagaaat tgtatttgtt
    10981 aaatatgtac tacaaactta gtagttggaa gggctaattc actcccaaag aagacaagat
    11041 atccttgatc tgtggatcta ccacacacaa ggctacttcc ctgattagca gaactacaca
    11101 ccagggccag gggtcagata tccactgacc tttggatggt gctacaagct agtaccagtt
    11161 gagccagata aggtagaaga ggccaataaa ggagagaaca ccagcttgtt acaccctgtg
    11221 agcctgcatg ggatggatga cccggagaga gaagtgttag agtggaggtt tgacagccgc
    11281 ctagcatttc atcacgtggc ccgagagctg catccggagt acttcaagaa ctgctgatat
    11341 cgagcttgct acaagggact ttccgctggg gactttccag ggaggcgtgg cctgggcggg
    11401 actggggagt ggcgagccct cagatcctgc atataagcag ctgctttttg cctgtactgg
    11461 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
    11521 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
    11581 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
    11641 tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg
    11701 actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca
    11761 aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag
    11821 cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa
    11881 tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct
    11941 ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt
    12001 cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg
    12061 catcaaagga tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa
    12121 aacaaaagta agaccaccgc acagcaagcg gccggccgct gatcttcaga cctggaggag
    12181 gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac
    12241 cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag
    12301 tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag
    12361 cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga
    12421 acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca
    12481 tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc
    12541 tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta
    12601 gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca
    12661 gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc
    12721 aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt
    12781 ttaacataac aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg
    12841 taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt
    12901 caccattatc gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa
    12961 tagaagaaga aggtggagag agagacagag acagatccat tcgattagtg aacggatctc
    13021 gacggtat
//

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