pLCRISPR-CMV

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pLCRISPRCMV载体基本信息

质粒名称: pLCRISPR-CMV 
出品公司: 爱迪基因
目录编号: 102609
存储实验室: Nicolas Manel

载体骨架基本信息
载体名称: pLCRISPR
载体出品公司: --
载体抗性: 氨苄青霉素
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: --
高拷贝/低拷贝: 高拷贝
生长条件: 30 ℃

插入基因信息
插入基因名称: --
物种来源: --
插入大小: --
基因突变: --
GenBank编号 --
启动子 --
标签/融合蛋白: --
克隆方法: --
5' 克隆位点: --
3' 克隆位点: --
5' 测序引物 --
3' 测序引物 --

文献引用方法
材料和方法部分: pLCRISPR-CMV was a gift from Nicolas Manel (Addgene plasmid # 102609 ; http://n2t.net/addgene:102609 ; RRID:Addgene_102609)
参考文献部分: Transmission of innate immune signaling by packaging of cGAMP in viral particles. Gentili M, Kowal J, Tkach M, Satoh T, Lahaye X, Conrad C, Boyron M, Lombard B, Durand S, Kroemer G, Loew D, Dalod M, Thery C, Manel N. Science. 2015 Sep 11;349(6253):1232-6. doi: 10.1126/science.aab3628. Epub 2015 Jul 30. 10.1126/science.aab3628 PubMed 26229115

pLCRISPRCMV载体图谱质粒图谱和多克隆位点信息

pLCRISPR-CMV质粒图谱

pLCRISPRCMV载体简介

pLCRISPRCMV载体序列

LOCUS       pLCRISPR-CMV               13834 bp ds-DNA     circular SYN 06-AUG-2018
DEFINITION  Control lentivector expressing Cas9 and gRNA scaffold.
ACCESSION   .
VERSION     .
KEYWORDS    pLCRISPR-CMV
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13834)
  AUTHORS   Gentili M, Kowal J, Tkach M, Satoh T, Lahaye X, Conrad C, Boyron M, 
            Lombard B, Durand S, Kroemer G, Loew D, Dalod M, Thery C, Manel N
  TITLE     Transmission of innate immune signaling by packaging of cGAMP in 
            viral particles.
  JOURNAL   Science. 2015 Sep 11;349(6253):1232-6. doi: 10.1126/science.aab3628.
            Epub 2015 Jul 30.
  PUBMED    26229115
REFERENCE   2  (bases 1 to 13834)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 6, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..13834
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(17..36)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        208..587
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        589..787
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             805..985
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1032..1157
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1650..1883
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     promoter        2367..2607
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     2367..2387
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2538..2557
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2634..2709
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    2743..2860
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2955..3258
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3259..3462
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      3516..3525
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             3525..3590
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3597..3617
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3642..7742
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7752..7799
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7809..7865
                     /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             7872..8471
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7872..7891)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     8368..8388
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8496..9084
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8549..8569)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8967..8978)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9087..9103)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9088..9104)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9609..9789
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9815..9832)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9821..10045
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10091..10519
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10178..10197)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10388..10409
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10528..10548)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10533..10862
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10713..10848
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10775..10794
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10910..10957
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10976..11350
                     /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    11480..11601
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11517..11536)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11571..11590
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11650..11666)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11650..11666)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11663..11685)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11674..11690
                     /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(11698..11728)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11743..11764
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11881..11898)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12052..12640)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12132..12151)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(12811..13671)
                     /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     13434..13453
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13672..13776)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatctgctcc
       61 ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct acaacaaggc
      121 aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt gcgctgcttc
      181 gcgatgtacg ggccagatat acgcgttgac attgattatt gactagttat taatagtaat
      241 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
      301 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      361 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      421 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
      481 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
      541 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
      601 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
      661 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
      721 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
      781 taagcagcgc gttttgcctg tactgggtct ctctggttag accagatctg agcctgggag
      841 ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc ttgagtgctt
      901 caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct cagacccttt
      961 tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa gcgaaaggga
     1021 aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg gcaagaggcg
     1081 aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag aaggagagag
     1141 atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg gaaaaaattc
     1201 ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg gcaagcaggg
     1261 agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc tgtagacaaa
     1321 tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga tcattatata
     1381 atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac accaaggaag
     1441 ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag caagcggccg
     1501 ctgatcttca gacctggagg aggagatatg agggacaatt ggagaagtga attatataaa
     1561 tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa gagaagagtg
     1621 gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt cttgggagca
     1681 gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg tacaggccag acaattattg
     1741 tctggtatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca acagcatctg
     1801 ttgcaactca cagtctgggg catcaagcag ctccaggcaa gaatcctggc tgtggaaaga
     1861 tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact catttgcacc
     1921 actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat ttggaatcac
     1981 acgacctgga tggagtggga cagagaaatt aacaattaca caagcttaat acactcctta
     2041 attgaagaat cgcaaaacca gcaagaaaag aatgaacaag aattattgga attagataaa
     2101 tgggcaagtt tgtggaattg gtttaacata acaaattggc tgtggtatat aaaattattc
     2161 ataatgatag taggaggctt ggtaggttta agaatagttt ttgctgtact ttctatagtg
     2221 aatagagtta ggcagggata ttcaccatta tcgtttcaga cccacctccc aaccccgagg
     2281 ggacccgaca ggcccgaagg aatagaagaa gaaggtggag agagagacag agacagatcc
     2341 attcgattag tgaacggatc ggcactgagg gcctatttcc catgattcct tcatatttgc
     2401 atatacgata caaggctgtt agagagataa ttggaattaa tttgactgta aacacaaaga
     2461 tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt gcagttttaa
     2521 aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct
     2581 tggctttata tatcttgtgg aaaggacgaa acaccgggtc ttcgagaaga cctgttttag
     2641 agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa agtggcaccg
     2701 agtcggtgct ttttttagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     2761 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     2821 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     2881 cagagatcca gtttggttaa ttaatagtaa tcaattacgg ggtcattagt tcatagccca
     2941 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
     3001 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
     3061 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
     3121 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
     3181 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
     3241 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
     3301 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
     3361 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
     3421 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag
     3481 atcccgcggg ctagcggatc cccaccggtc gtacggccac catggactat aaggaccacg
     3541 acggagacta caaggatcat gatattgatt acaaagacga tgacgataag atggccccaa
     3601 agaagaagcg gaaggtcggt atccacggag tcccagcagc cgacaagaag tacagcatcg
     3661 gcctggacat cggcaccaac tctgtgggct gggccgtgat caccgacgag tacaaggtgc
     3721 ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca cagcatcaag aagaacctga
     3781 tcggagccct gctgttcgac agcggcgaaa cagccgaggc cacccggctg aagagaaccg
     3841 ccagaagaag atacaccaga cggaagaacc ggatctgcta tctgcaagag atcttcagca
     3901 acgagatggc caaggtggac gacagcttct tccacagact ggaagagtcc ttcctggtgg
     3961 aagaggataa gaagcacgag cggcacccca tcttcggcaa catcgtggac gaggtggcct
     4021 accacgagaa gtaccccacc atctaccacc tgagaaagaa actggtggac agcaccgaca
     4081 aggccgacct gcggctgatc tatctggccc tggcccacat gatcaagttc cggggccact
     4141 tcctgatcga gggcgacctg aaccccgaca acagcgacgt ggacaagctg ttcatccagc
     4201 tggtgcagac ctacaaccag ctgttcgagg aaaaccccat caacgccagc ggcgtggacg
     4261 ccaaggccat cctgtctgcc agactgagca agagcagacg gctggaaaat ctgatcgccc
     4321 agctgcccgg cgagaagaag aatggcctgt tcggcaacct gattgccctg agcctgggcc
     4381 tgacccccaa cttcaagagc aacttcgacc tggccgagga tgccaaactg cagctgagca
     4441 aggacaccta cgacgacgac ctggacaacc tgctggccca gatcggcgac cagtacgccg
     4501 acctgtttct ggccgccaag aacctgtccg acgccatcct gctgagcgac atcctgagag
     4561 tgaacaccga gatcaccaag gcccccctga gcgcctctat gatcaagaga tacgacgagc
     4621 accaccagga cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca
     4681 aagagatttt cttcgaccag agcaagaacg gctacgccgg ctacattgac ggcggagcca
     4741 gccaggaaga gttctacaag ttcatcaagc ccatcctgga aaagatggac ggcaccgagg
     4801 aactgctcgt gaagctgaac agagaggacc tgctgcggaa gcagcggacc ttcgacaacg
     4861 gcagcatccc ccaccagatc cacctgggag agctgcacgc cattctgcgg cggcaggaag
     4921 atttttaccc attcctgaag gacaaccggg aaaagatcga gaagatcctg accttccgca
     4981 tcccctacta cgtgggccct ctggccaggg gaaacagcag attcgcctgg atgaccagaa
     5041 agagcgagga aaccatcacc ccctggaact tcgaggaagt ggtggacaag ggcgcttccg
     5101 cccagagctt catcgagcgg atgaccaact tcgataagaa cctgcccaac gagaaggtgc
     5161 tgcccaagca cagcctgctg tacgagtact tcaccgtgta taacgagctg accaaagtga
     5221 aatacgtgac cgagggaatg agaaagcccg ccttcctgag cggcgagcag aaaaaggcca
     5281 tcgtggacct gctgttcaag accaaccgga aagtgaccgt gaagcagctg aaagaggact
     5341 acttcaagaa aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa gatcggttca
     5401 acgcctccct gggcacatac cacgatctgc tgaaaattat caaggacaag gacttcctgg
     5461 acaatgagga aaacgaggac attctggaag atatcgtgct gaccctgaca ctgtttgagg
     5521 acagagagat gatcgaggaa cggctgaaaa cctatgccca cctgttcgac gacaaagtga
     5581 tgaagcagct gaagcggcgg agatacaccg gctggggcag gctgagccgg aagctgatca
     5641 acggcatccg ggacaagcag tccggcaaga caatcctgga tttcctgaag tccgacggct
     5701 tcgccaacag aaacttcatg cagctgatcc acgacgacag cctgaccttt aaagaggaca
     5761 tccagaaagc ccaggtgtcc ggccagggcg atagcctgca cgagcacatt gccaatctgg
     5821 ccggcagccc cgccattaag aagggcatcc tgcagacagt gaaggtggtg gacgagctcg
     5881 tgaaagtgat gggccggcac aagcccgaga acatcgtgat cgaaatggcc agagagaacc
     5941 agaccaccca gaagggacag aagaacagcc gcgagagaat gaagcggatc gaagagggca
     6001 tcaaagagct gggcagccag atcctgaaag aacaccccgt ggaaaacacc cagctgcaga
     6061 acgagaagct gtacctgtac tacctgcaga atgggcggga tatgtacgtg gaccaggaac
     6121 tggacatcaa ccggctgtcc gactacgatg tggaccatat cgtgcctcag agctttctga
     6181 aggacgactc catcgacaac aaggtgctga ccagaagcga caagaaccgg ggcaagagcg
     6241 acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg cagctgctga
     6301 acgccaagct gattacccag agaaagttcg acaatctgac caaggccgag agaggcggcc
     6361 tgagcgaact ggataaggcc ggcttcatca agagacagct ggtggaaacc cggcagatca
     6421 caaagcacgt ggcacagatc ctggactccc ggatgaacac taagtacgac gagaatgaca
     6481 agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga
     6541 aggatttcca gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct
     6601 acctgaacgc cgtcgtggga accgccctga tcaaaaagta ccctaagctg gaaagcgagt
     6661 tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat gatcgccaag agcgagcagg
     6721 aaatcggcaa ggctaccgcc aagtacttct tctacagcaa catcatgaac tttttcaaga
     6781 ccgagattac cctggccaac ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg
     6841 aaaccgggga gatcgtgtgg gataagggcc gggattttgc caccgtgcgg aaagtgctga
     6901 gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag
     6961 agtctatcct gcccaagagg aacagcgata agctgatcgc cagaaagaag gactgggacc
     7021 ctaagaagta cggcggcttc gacagcccca ccgtggccta ttctgtgctg gtggtggcca
     7081 aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa agagctgctg gggatcacca
     7141 tcatggaaag aagcagcttc gagaagaatc ccatcgactt tctggaagcc aagggctaca
     7201 aagaagtgaa aaaggacctg atcatcaagc tgcctaagta ctccctgttc gagctggaaa
     7261 acggccggaa gagaatgctg gcctctgccg gcgaactgca gaagggaaac gaactggccc
     7321 tgccctccaa atatgtgaac ttcctgtacc tggccagcca ctatgagaag ctgaagggct
     7381 cccccgagga taatgagcag aaacagctgt ttgtggaaca gcacaagcac tacctggacg
     7441 agatcatcga gcagatcagc gagttctcca agagagtgat cctggccgac gctaatctgg
     7501 acaaagtgct gtccgcctac aacaagcacc gggataagcc catcagagag caggccgaga
     7561 atatcatcca cctgtttacc ctgaccaatc tgggagcccc tgccgccttc aagtactttg
     7621 acaccaccat cgaccggaag aggtacacca gcaccaaaga ggtgctggac gccaccctga
     7681 tccaccagag catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggcg
     7741 acgcggccgc caagcgtcct gctgctacta agaaagctgg tcaagctaag aaaaagaaag
     7801 gaagcggagc tactaacttc agcctgctga agcaggctgg agacgtggag gagaaccctg
     7861 gacctggatc catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc
     7921 ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg
     7981 tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg
     8041 tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga
     8101 ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg
     8161 agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc
     8221 ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca
     8281 agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc
     8341 ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca
     8401 ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc
     8461 ccggtgccta agaattcgat atcaagctta tcgataatca acctctggat tacaaaattt
     8521 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     8581 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     8641 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     8701 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     8761 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     8821 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     8881 tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc
     8941 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     9001 gcctgctgcc ggctctgcgg cctcttccgc atcttcgcct tcgccctcag acgagtcgga
     9061 tctccctttg ggccgcctcc ccgcatcgat accgtcgacc tcgagaccta gaaaaacatg
     9121 gagcaatcac aagtagcaat acagcagcta ccaatgctga ttgtgcctgg ctagaagcac
     9181 aagaggagga ggaggtgggt tttccagtca cacctcaggt acctttaaga ccaatgactt
     9241 acaaggcagc tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa
     9301 ttcactccca acgaagtcaa gatatccttg atctgtggat ctaccacaca caaggctact
     9361 tccctgattg gcagaactac acaccagggc cagggatcag atatccactg acctttggat
     9421 ggtgctacaa gctagtacca gttgagcaag agaaggtaga agaagccaat gaaggagaga
     9481 acacccgctt gttacaccct gtgagcctgc atgggatgga tgacccggag agagaagtat
     9541 tagagtggag gtttgacagc cgcctagcat ttcatcacat ggcccgagag ctgcatccgg
     9601 actgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     9661 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     9721 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     9781 tctctagcag ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag
     9841 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     9901 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     9961 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagataa tagcaggcat
    10021 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg
    10081 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
    10141 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
    10201 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
    10261 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
    10321 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
    10381 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
    10441 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
    10501 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc
    10561 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
    10621 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
    10681 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
    10741 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
    10801 ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
    10861 aaaagctccc gggagcttgt atatccattt tcggatctga tcagcacgtg ttgacaatta
    10921 atcatcggca tagtatatcg gcatagtata atacgacaag gtgaggaact aaaccatggc
    10981 caagttgacc agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt
    11041 ctggaccgac cggctcgggt tctcccggga cttcgtggag gacgacttcg ccggtgtggt
    11101 ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc cggacaacac
    11161 cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt cggaggtcgt
    11221 gtccacgaac ttccgggacg cctccgggcc ggccatgacc gagatcggcg agcagccgtg
    11281 ggggcgggag ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg tggccgagga
    11341 gcaggactga cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg
    11401 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
    11461 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
    11521 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
    11581 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
    11641 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
    11701 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
    11761 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
    11821 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
    11881 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
    11941 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
    12001 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
    12061 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
    12121 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
    12181 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
    12241 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
    12301 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
    12361 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
    12421 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
    12481 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
    12541 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
    12601 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
    12661 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
    12721 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
    12781 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
    12841 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
    12901 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
    12961 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
    13021 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
    13081 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
    13141 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
    13201 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
    13261 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
    13321 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
    13381 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
    13441 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
    13501 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
    13561 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
    13621 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
    13681 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
    13741 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
    13801 tgacgtcgac ggatcgggag atctcccgat cccc
//

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