质粒名称: | pMGS56(GFP-ARF16-PB1-P2A-OsTIR1) |
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出品公司: | addgene |
目录编号: | 129668 |
存储实验室: | Michael Guertin |
载体骨架基本信息 |
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载体名称: | pMK232 |
载体出品公司: | Masato Kanemaki |
空载体大小: | 9590 bp |
完整质粒大小: | 10961 bp |
载体类型: | 哺乳细胞表达 |
筛选标记: | Puromycin |
载体抗性: | 氨苄霉素 |
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: | OsARF16-PB1 domain |
物种来源: | Oryza sativa |
插入大小: | 1377bp |
标签/融合蛋白: | GFP (N terminal on insert) |
克隆方法: | Ligation Independent Cloning |
5’测序引物 | EGFP-C |
文献引用方法 |
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材料和方法部分: | pMGS56 (GFP-ARF16-PB1-P2A-OsTIR1) was a gift from Michael Guertin (Addgene plasmid # 129668 ; http://n2t.net/addgene:129668 ; RRID:Addgene_129668) |
参考文献部分: | An improved auxin-inducible degron system preserves native protein levels and enables rapid and specific protein depletion. Sathyan KM, McKenna BD, Anderson WD, Duarte FM, Core L, Guertin MJ. Genes Dev. 2019 Oct 1;33(19-20):1441-1455. doi: 10.1101/gad.328237.119. Epub 2019 Aug 29. 10.1101/gad.328237.119 PubMed 31467088 |
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载体基本信息
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载体简介
载体序列
LOCUS pMGS56(GFP-ARF16-PB1-P2A-OsTIR1) 10961 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION A repair construct to express GFP-ARF16-PB1 and OsTIR1 under the control of the CMV promoter from the human AAVS1 locus. ACCESSION . VERSION . KEYWORDS pMGS56 (GFP-ARF16-PB1-P2A-OsTIR1) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 10961) AUTHORS Sathyan KM, McKenna BD, Anderson WD, Duarte FM, Core L, Guertin MJ TITLE An improved auxin-inducible degron system preserves native protein levels and enables rapid and specific protein depletion. JOURNAL Genes Dev. 2019 Oct 1;33(19-20):1441-1455. doi: 10.1101/gad.328237.119. Epub 2019 Aug 29. PUBMED 31467088 REFERENCE 2 (bases 1 to 10961) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.biofeng.com FEATURES Location/Qualifiers source 1..10961 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind 107..128 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 143..173 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 181..197 /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)." primer_bind 186..208 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 205..221 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 205..221 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" promoter 242..260 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" misc_feature 295..1098 /label=HA-L /note="left homology arm from the adeno-associated virus integration site (AAVS1) within intron 1 of the human PPP1R12C gene" enhancer 1155..1458 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1459..1662 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 1613..1633 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 1694..1703 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1700..2416 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 1700..2416 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1745..1766) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2006..2025) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2353..2374 /label=EGFP-C /note="EGFP, forward primer" CDS 2444..2464 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 3011..3067 /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 3077..4804 /codon_start=1 /product="rice transport inhibitor response 1 protein" /label=OsTIR1 /note="F-box protein that mediates auxin-dependent interaction of a ubiquitin ligase with an auxin-inducible degron (Tan et al., 2007)" /translation="MTYFPEEVVEHIFSFLPAQRDRNTVSLVCKVWYEIERLSRRGVFV GNCYAVRAGRVAARFPNVRALTVKGKPHFADFNLVPPDWGGYAGPWIEAAARGCHGLEE LRMKRMVVSDESLELLARSFPRFRALVLISCEGFSTDGLAAVASHCKLLRELDLQENEV EDRGPRWLSCFPDSCTSLVSLNFACIKGEVNAGSLERLVSRSPNLRSLRLNRSVSVDTL AKILLRTPNLEDLGTGNLTDDFQTESYFKLTSALEKCKMLRSLSGFWDASPVCLSFIYP LCAQLTGLNLSYAPTLDASDLTKMISRCVKLQRLWVLDCISDKGLQVVASSCKDLQELR VFPSDFYVAGYSAVTEEGLVAVSLGCPKLNSLLYFCHQMTNAALVTVAKNCPNFTRFRL CILEPGKPDVVTSQPLDEGFGAIVRECKGLQRLSISGLLTDKVFMYIGKYAKQLEMLSI AFAGDSDKGMMHVMNGCKNLRKLEIRDSPFGDAALLGNFARYETMRSLWMSSCNVTLKG CQVLASKMPMLNVEVINERDGSNEMEENHGDLPKVEKLYVYRTTAGARDDAPNFVKIL" polyA_signal 4978..5099 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5015..5034) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5069..5088 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" promoter 5108..5608 /label=hPGK promoter /note="human phosphoglycerate kinase 1 promoter" primer_bind 5527..5545 /label=hPGK-F /note="Human PGK promoter, forward primer" CDS 5630..6229 /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(5630..5649) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 6126..6146 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 6236..6460 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" misc_feature 6463..7299 /label=HA-R /note="right homology arm from the adeno-associated virus integration site (AAVS1) within intron 1 of the human PPP1R12C gene" primer_bind complement(7333..7352) /label=T7 /note="T7 promoter, forward primer" promoter complement(7334..7352) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(7359..7376) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(7359..7375) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(7368..7390) /label=M13/pUC Forward /note="In lacZ gene" CDS 7513..7815 /codon_start=1 /gene="ccdB" /product="CcdB, a bacterial toxin that poisons DNA gyrase" /label=ccdB /note="Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains." /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" primer_bind 7694..7713 /label=ccdB-fwd /note="ccdB gene, forward primer" CDS 8164..8958 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(8218..8237) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 8828..8847 /label=Neo-F /note="Neomycin resistance gene, forward primer" CDS complement(9208..10068) /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 9831..9850 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 10192..10780 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 10681..10700 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 10934..10951 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc 61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc 121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa 181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctca 241 gaattaaccc tcactaaagg gactagtcct gcaggtttaa acgaattcgc cctttgcttt 301 ctctgaccag cattctctcc cctgggcctg tgccgctttc tgtctgcagc ttgtggcctg 361 ggtcacctct acggctggcc cagatccttc cctgccgcct ccttcaggtt ccgtcttcct 421 ccactccctc ttccccttgc tctctgctgt gttgctgccc aaggatgctc tttccggagc 481 acttccttct cggcgctgca ccacgtgatg tcctctgagc ggatcctccc cgtgtctggg 541 tcctctccgg gcatctctcc tccctcaccc aaccccatgc cgtcttcact cgctgggttc 601 ccttttcctt ctccttctgg ggcctgtgcc atctctcgtt tcttaggatg gccttctccg 661 acggatgtct cccttgcgtc ccgcctcccc ttcttgtagg cctgcatcat caccgttttt 721 ctggacaacc ccaaagtacc ccgtctccct ggctttagcc acctctccat cctcttgctt 781 tctttgcctg gacaccccgt tctcctgtgg attcgggtca cctctcactc ctttcatttg 841 ggcagctccc ctacccccct tacctctcta gtctgtgcta gctcttccag ccccctgtca 901 tggcatcttc caggggtccg agagctcagc tagtcttctt cctccaaccc gggcccctat 961 gtccacttca ggacagcatg tttgctgcct ccagggatcc tgtgtccccg agctgggacc 1021 accttatatt cccagggccg gttaatgtgg ctctggttct gggtactttt atctgtcccc 1081 tccaccccac agtggggcaa ttaatagtaa tcaattacgg ggtcattagt tcatagccca 1141 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac 1201 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact 1261 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa 1321 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg 1381 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta 1441 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg 1501 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg 1561 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg 1621 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag 1681 atccgctagc gctgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc 1741 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg 1801 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct 1861 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg 1921 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt 1981 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa 2041 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga 2101 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat 2161 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga 2221 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt 2281 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga 2341 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat 2401 ggacgagctg tacaagtccg gccggactca gatctcgagc atgcctaaga agaagcggaa 2461 ggtgggcagc acactgactt ccgggcagcg ggatcaccgg gacgtcgaga acgagctgag 2521 ctccgccgcc ttcagctccc agagcttcgg ggtgcccgac atgagcttta agcccggctg 2581 cagcagcgac gtggccgtga ccgacgccgg gatgcccagc caggggctgt ggaacaacca 2641 gacccagcgg atgaggacat tcacaaaggt gcagaagcgg gggagcgtgg gccggagcat 2701 cgatatcacc aggtacaggg attacgatga gctgaggcat gatctcgcct gtatgttcgg 2761 catccagggc cagctggagg acccctaccg gatggactgg aagctggtgt acgtggacca 2821 cgagaacgat atcctgctgg tgggcgatga cccctgggag gagttcgtgg gctgcgtgaa 2881 gagcattaag attctgtctg ctgctgaggt gcagcagatg agcctggatg gagatctggg 2941 aggagtgcca cctcagactc aggcctgtag tgctagtgat gatgctaacg cctggagagg 3001 gggaagcgga gctactaact tcagcctgct gaagcaggct ggagacgtgg aggagaaccc 3061 tggacctgct gccaccatga catactttcc tgaagaggtc gtcgaacaca tttttagctt 3121 cctgcctgca cagagagata gaaacacagt gagcctggtc tgcaaagtgt ggtacgagat 3181 cgaacgcctg agccggagag gagtgttcgt cggcaactgc tatgctgtga gagcaggcag 3241 ggtcgccgct aggtttccaa atgtgcgcgc actgaccgtc aaggggaaac cccacttcgc 3301 cgactttaac ctggtgcccc ctgattgggg aggatacgcc ggcccttgga tcgaggcagc 3361 cgctcgcggc tgtcatggac tggaggaact gcgcatgaag cgaatggtgg tctctgacga 3421 aagtctggag ctgctggctc ggagcttccc taggtttcgc gcactggtgc tgatttcttg 3481 cgaaggcttc agcaccgatg gactggcagc cgtggcctcc cactgtaagc tgctgcggga 3541 gctggacctc caggagaatg aagtggagga tagaggcccc agatggctgt cttgcttccc 3601 agactcatgt accagcctgg tgtccctgaa ctttgcctgc atcaaaggcg aagtgaatgc 3661 tgggtccctg gagcggctgg tctcaagaag ccccaacctg aggtctctgc ggctgaaccg 3721 gagcgtgagc gtggacactc tggctaagat tctgctgaga acccctaacc tggaggatct 3781 gggaaccggc aatctgacag acgatttcca gacagaatcc tactttaaac tgacttctgc 3841 cctggagaag tgtaaaatgc tgaggagtct gtcaggattc tgggatgctt cacccgtgtg 3901 cctgagcttt atctaccctc tgtgtgcaca gctgacaggc ctgaacctga gctatgcacc 3961 aaccctggac gccagtgatc tgacaaagat gatctcacgc tgcgtgaaac tccagcgact 4021 gtgggtgctg gactgtattt ccgataaggg gctccaggtg gtcgccagct cctgcaagga 4081 cctccaggag ctgagagtgt tcccatctga tttttacgtg gccggatata gtgctgtcac 4141 tgaggaaggc ctggtggcag tctcactggg atgcccaaag ctgaacagcc tgctgtattt 4201 ctgtcatcag atgactaatg ctgcactggt gaccgtcgcc aagaactgcc ctaatttcac 4261 ccgatttcgg ctgtgtattc tggaaccagg caaacccgac gtggtcacat cccagccact 4321 ggatgaaggg tttggagcta tcgtgagaga gtgcaaggga ctccagaggc tgagcatttc 4381 cggcctgctg acagacaaag tgttcatgta catcggcaag tatgctaagc agctggagat 4441 gctgagcatt gcatttgccg gagactccga taagggcatg atgcacgtga tgaacgggtg 4501 taagaatctg cgaaaactgg aaatccggga cagccctttc ggggatgccg ctctgctggg 4561 aaactttgcc agatacgaga caatgaggag cctgtggatg tctagttgca atgtgactct 4621 gaagggctgt caggtcctgg ctagtaaaat gcctatgctg aacgtggaag tcattaatga 4681 gcgggacggg tctaacgaaa tggaggaaaa tcatggcgac ctgccaaagg tggagaaact 4741 gtatgtgtat cggaccaccg caggggcaag agatgatgct cccaactttg tgaagattct 4801 gtgaggatcc atcggatccc gggcccgtcg acggtaccgc gggcccggga tccaccgggg 4861 ccgcgactct agatcataat cagccatacc acatttgtag aggttttact tgctttaaaa 4921 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac 4981 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat 5041 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa 5101 gttggggttg cgccttttcc aaggcagccc tgggtttgcg cagggacgcg gctgctctgg 5161 gcgtggttcc gggaaacgca gcggcgccga ccctgggtct cgcacattct tcacgtccgt 5221 tcgcagcgtc acccggatct tcgccgctac ccttgtgggc cccccggcga cgcttcctgc 5281 tccgccccta agtcgggaag gttccttgcg gttcgcggcg tgccggacgt gacaaacgga 5341 agccgcacgt ctcactagta ccctcgcaga cggacagcgc cagggagcaa tggcagcgcg 5401 ccgaccgcga tgggctgtgg ccaatagcgg ctgctcagca gggcgcgccg agagcagcgg 5461 ccgggaaggg gcggtgcggg aggcggggtg tggggcggta gtgtgggccc tgttcctgcc 5521 cgcgcggtgt tccgcattct gcaagcctcc ggagcgcacg tcggcagtcg gctccctcgt 5581 tgaccgaatc accgacctct ctccccaggg ggatccaccg gagcttacca tgaccgagta 5641 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc 5701 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga 5761 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt 5821 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc 5881 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc 5941 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt 6001 cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt 6061 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc 6121 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt 6181 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgag gtaccctgtg 6241 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa 6301 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt 6361 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa 6421 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg ttactaggga caggattggt 6481 gacagaaaag ccccatcctt aggcctcctc cttcctagtc tcctgatatt gggtctaacc 6541 cccacctcct gttaggcaga ttccttatct ggtgacacac ccccatttcc tggagccatc 6601 tctctccttg ccagaacctc taaggtttgc ttacgatgga gccagagagg atcctgggag 6661 ggagagcttg gcagggggtg ggagggaagg gggggatgcg tgacctgccc ggttctcagt 6721 ggccaccctg cgctaccctc tcccagaacc tgagctgctc tgacgcggct gtctggtgcg 6781 tttcactgat cctggtgctg cagcttcctt acacttccca agaggagaag cagtttggaa 6841 aaacaaaatc agaataagtt ggtcctgagt tctaactttg gctcttcacc tttctagtcc 6901 ccaatttata ttgttcctcc gtgcgtcagt tttacctgtg agataaggcc agtagccagc 6961 cccgtcctgg cagggctgtg gtgaggaggg gggtgtccgt gtggaaaact ccctttgtga 7021 gaatggtgcg tcctaggtgt tcaccaggtc gtggccgcct ctactccctt tctctttctc 7081 catccttctt tccttaaaga gtccccagtg ctatctggga catattcctc cgcccagagc 7141 agggtcccgc ttccctaagg ccctgctctg ggcttctggg tttgagtcct tggcaagccc 7201 aggagaggcg ctcaggcttc cctgtccccc ttcctcgtcc accatctcat gcccctggct 7261 ctcctgcccc ttccctacag gggttcctgg ctctgctcta agggcgaatt cgcggccgct 7321 aaattcaatt cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt 7381 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc 7441 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc 7501 ctatacgtac ggcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt 7561 gtggatgtac agagtgatat tattgacacg ccggggcgac ggatggtgat ccccctggcc 7621 agtgcacgtc tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg 7681 gatgaaagct ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg 7741 gaagaagtgg ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg 7801 ttctggggaa tataaatgtc aggcatgaga ttatcaaaaa ggatcttcac ctagatcctt 7861 ttcacgtaga aagccagtcc gcagaaacgg tgctgacccc ggatgaatgt cagctactgg 7921 gctatctgga caagggaaaa cgcaagcgca aagagaaagc aggtagcttg cagtgggctt 7981 acatggcgat agctagactg ggcggtttta tggacagcaa gcgaaccgga attgccagct 8041 ggggcgccct ctggtaaggt tgggaagccc tgcaaagtaa actggatggc tttcttgccg 8101 ccaaggatct gatggcgcag gggatcaagc tctgatcaag agacaggatg aggatcgttt 8161 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta 8221 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg 8281 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa 8341 ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct 8401 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg 8461 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca 8521 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat 8581 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac 8641 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc 8701 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa 8761 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag 8821 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc 8881 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt 8941 cttgacgagt tcttctgaat tattaacgct tacaatttcc tgatgcggta ttttctcctt 9001 acgcatctgt gcggtatttc acaccgcatc aggtggcact tttcggggaa atgtgcgcgg 9061 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagattatc 9121 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag 9181 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc 9241 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac 9301 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc 9361 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg 9421 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag 9481 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc 9541 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac 9601 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag 9661 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac 9721 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg 9781 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc 9841 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact 9901 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg 9961 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa 10021 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt 10081 tcaatattat tgaagcattt atcagggtta ttgtctcatg accaaaatcc cttaacgtga 10141 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc 10201 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt 10261 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc 10321 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc 10381 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg 10441 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg 10501 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga 10561 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc 10621 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg 10681 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg 10741 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt 10801 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc 10861 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg 10921 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa g //
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