质粒名称: | pINDUCER21 (ORF-EG) |
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出品公司: | 爱迪基因 |
目录编号: | 46948 |
存储实验室: | Stephen Elledge, Thomas Westbrook |
载体骨架基本信息 |
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载体名称: | pHAGE-CAGS-W |
载体出品公司: | Dr. Richard Mulligan |
载体抗性: | 氯霉素和氨苄青霉素 |
空载体大小: | 4874 bp |
完整质粒大小: | -- |
载体修饰: | pINDUCER21(ORF-UG)是用NdeI和SphI消化pINDUCER20制成的。eGFP分别在5′端和3′端用AgeI和SphI进行PCR扩增,然后用AgeI和SphI进行酶切。将载体与eGFP连接,并进行序列验证。pINDUCER20(ORF-UN)是通过用BamHI和SpeI从pSLIK Neo-TGmiR-Luc载体中消化TRE2,并将钝端克隆到SpeI和XhoI消化的噬菌体-CAGS-W载体(Richard Mulligan博士,哈佛医学院,波士顿)的骨干中来产生噬菌体TRE载体。用SpeI从噬菌体EF-DEST载体(G.H.)中切出含有网关受体盒的片段并插入噬菌体TRE中以产生噬菌体TRE-DEST载体。用PacI和ClaI从pSLIK-Neo-TGmiR-Luc载体中切出Ubc-rtTA3-IRES-Neo片段,并将其克隆到SphI和ClaI消化的噬菌体tredest的骨架中。 |
载体类型: | 慢病毒载体表达 |
筛选标记: | eGFP |
高拷贝/低拷贝: | -- |
生长条件: | 30 ℃ |
插入基因信息 |
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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 |
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材料和方法部分: | pINDUCER21 (ORF-EG) was a gift from Stephen Elledge & Thomas Westbrook (Addgene plasmid # 46948 ; http://n2t.net/addgene:46948 ; RRID:Addgene_46948) |
参考文献部分: | The pINDUCER lentiviral toolkit for inducible RNA interference in vitro and in vivo. Meerbrey KL, Hu G, Kessler JD, Roarty K, Li MZ, Fang JE, Herschkowitz JI, Burrows AE, Ciccia A, Sun T, Schmitt EM, Bernardi RJ, Fu X, Bland CS, Cooper TA, Schiff R, Rosen JM, Westbrook TF, Elledge SJ. Proc Natl Acad Sci U S A. 2011 Mar 1;108(9):3665-70. doi: 10.1073/pnas.1019736108. Epub 2011 Feb 9. 10.1073/pnas.1019736108 PubMed 21307310 |
买家导航
载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS pINDUCER21 (ORF-EG) 12152 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Inducible lentiviral gene expression vector. Chloramphenicol cm + ccdB cassette may be removed to clone in gene of interest. ACCESSION . VERSION . KEYWORDS pINDUCER21 (ORF-EG) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 12152) AUTHORS Meerbrey KL, Hu G, Kessler JD, Roarty K, Li MZ, Fang JE, Herschkowitz JI, Burrows AE, Ciccia A, Sun T, Schmitt EM, Bernardi RJ, Fu X, Bland CS, Cooper TA, Schiff R, Rosen JM, Westbrook TF, Elledge SJ TITLE The pINDUCER lentiviral toolkit for inducible RNA interference in vitro and in vivo. JOURNAL Proc Natl Acad Sci U S A. 2011 Mar 1;108(9):3665-70. doi: 10.1073/pnas.1019736108. Epub 2011 Feb 9. PUBMED 21307310 REFERENCE 2 (bases 1 to 12152) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..12152 /organism="synthetic DNA construct" /mol_type="other DNA" LTR 1..634 /label=3' LTR /note="3' long terminal repeat (LTR) from HIV-1" misc_feature 681..806 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1303..1536 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 1721..1765 /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" misc_feature 2058..2175 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" protein_bind 2230..2500 /label=tetracycline response element /bound_moiety="tetracycline repressor TetR" /note="contains seven copies of the tetracycline operator tetO" promoter 2533..2571 /label=minimal CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 2568..2592 /label=LNCX /note="Human CMV promoter, forward primer" protein_bind 2701..2825 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 2850..2880 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 2934..3593 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind complement(3000..3019) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 3935..4240 /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="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" primer_bind 4119..4138 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(4281..4405) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" CDS 4458..4484 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" promoter 4704..5882 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 4935..5873 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 5830..5850 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" protein_bind complement(5952..5985) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS 5992..6738 /codon_start=1 /product="improved tetracycline-controlled transactivator" /label=rtTA-Advanced /note="In the Tet-On(R) system, rtTA-Advanced binds to the Tet-responsive element and stimulates transcription only in the presence of tetracycline or doxycycline." /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA DALDDFDLDMLPADALDDFDLDMLPG" misc_feature 6791..7364 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(6958..6975) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 7185..7204 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 7380..8099 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 7380..8096 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(7425..7446) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(7686..7705) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 8033..8054 /label=EGFP-C /note="EGFP, forward primer" primer_bind 8235..8254 /label=Neo-F /note="Neomycin resistance gene, forward primer" misc_feature 8542..9130 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(8595..8615) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(9013..9024) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 9205..9438 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" primer_bind complement(9557..9575) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 9643..9747 /gene="bla" /label=AmpR promoter CDS 9748..10608 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(9966..9985) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 10779..11367 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 11268..11287 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 11521..11538 /label=L4440 /note="L4440 vector, forward primer" primer_bind 11630..11649 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" protein_bind 11778..11799 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 11814..11844 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 11852..11868 /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 11857..11879 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 11876..11892 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 11876..11892 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind 11942..11961 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" ORIGIN 1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca 61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac 121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca 181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg 241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag 301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg 361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat 421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga 481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct 541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc 601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag 661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg 721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga 781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg 841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg 901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct 961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat 1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca 1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc 1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag 1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc 1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg 1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc 1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc 1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct 1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa 1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca 1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt 1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt 1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta 1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt 1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct 1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga 1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccgaa ttcacaaatg 2041 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa 2101 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca 2161 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gactaggatc 2221 ctttaccact ccctatcagt gatagagaaa agtgaaagtc gagtttacca ctccctatca 2281 gtgatagaga aaagtgaaag tcgagtttac cactccctat cagtgataga gaaaagtgaa 2341 agtcgagttt accactccct atcagtgata gagaaaagtg aaagtcgagt ttaccactcc 2401 ctatcagtga tagagaaaag tgaaagtcga gtttaccact ccctatcagt gatagagaaa 2461 agtgaaagtc gagtttacca ctccctatca gtgatagaga aaagtgaaag tcgagctcgg 2521 tacccgggtc gaggtaggcg tgtacggtgg gaggcctata taagcagagc tcgtttagtg 2581 aaccgtcaga tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg 2641 gaccgatcca gcctccgcgg ccccgaacta gtccagtgtg gtggaattct gcagatatca 2701 acaagtttgt acaaaaaagc tgaacgagaa acgtaaaatg atataaatat caatatatta 2761 aattagattt tgcataaaaa acagactaca taatactgta aaacacaaca tatccagtca 2821 ctatggcggc cgcattaggc accccaggct ttacacttta tgcttccggc tcgtataatg 2881 tgtggatttt gagttaggat ccgtcgagat tttcaggagc taaggaagct aaaatggaga 2941 aaaaaatcac tggatatacc accgttgata tatcccaatg gcatcgtaaa gaacattttg 3001 aggcatttca gtcagttgct caatgtacct ataaccagac cgttcagctg gatattacgg 3061 cctttttaaa gaccgtaaag aaaaataagc acaagtttta tccggccttt attcacattc 3121 ttgcccgcct gatgaatgct catccggaat tccgtatggc aatgaaagac ggtgagctgg 3181 tgatatggga tagtgttcac ccttgttaca ccgttttcca tgagcaaact gaaacgtttt 3241 catcgctctg gagtgaatac cacgacgatt tccggcagtt tctacacata tattcgcaag 3301 atgtggcgtg ttacggtgaa aacctggcct atttccctaa agggtttatt gagaatatgt 3361 ttttcgtctc agccaatccc tgggtgagtt tcaccagttt tgatttaaac gtggccaata 3421 tggacaactt cttcgccccc gttttcacca tgggcaaata ttatacgcaa ggcgacaagg 3481 tgctgatgcc gctggcgatt caggttcatc atgccgtttg tgatggcttc catgtcggca 3541 gaatgcttaa tgaattacaa cagtactgcg atgagtggca gggcggggcg taaagatctg 3601 gatccggctt actaaaagcc agataacagt atgcgtattt gcgcgctgat ttttgcggta 3661 taagaatata tactgatatg tatacccgaa gtatgtcaaa aagaggtatg ctatgaagca 3721 gcgtattaca gtgacagttg acagcgacag ctatcagttg ctcaaggcat atatgatgtc 3781 aatatctccg gtctggtaag cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa 3841 cgctggaaag cggaaaatca ggaagggatg gctgaggtcg cccggtttat tgaaatgaac 3901 ggctcttttg ctgacgagaa caggggctgg tgaaatgcag tttaaggttt acacctataa 3961 aagagagagc cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgcccgg 4021 gcgacggatg gtgatccccc tggccagtgc acgtctgctg tcagataaag tctcccgtga 4081 actttacccg gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc 4141 cagtgtgccg gtctccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga 4201 catcaaaaac gccattaacc tgatgttctg gggaatataa atgtcaggct cccttataca 4261 cagccagtct gcaggtcgac catagtgact ggatatgttg tgttttacag tattatgtag 4321 tctgtttttt atgcaaaatc taatttaata tattgatatt tatatcattt tacgtttctc 4381 gttcagcttt cttgtacaaa gtggttgata tccagcacag tggcggccgc tcgagtctag 4441 agggcccgcg gttcgaatac ccatacgacg tcccagacta cgcttagtaa tgattaatta 4501 aactagaaat tctaccgggt aggggaggcg cttttcccaa ggcagtctgg agtaacccac 4561 ccaagatctg gcctccgcgc cgggttttgg cgcctcccgc gggcgccccc ctcctcacgg 4621 cgagccgcgt aagatctggc ctccgcgccg ggttttggcg cctcccgcgg gcgcccccct 4681 cctcacggcg agcctagtcg tgaggctccg gtgcccgtca gtgggcagag cgcacatcgc 4741 ccacagtccc cgagaagttg gggggagggg tcggcaattg aaccggtgcc tagagaaggt 4801 ggcgcggggt aaactgggaa agtgatgtcg tgtactggct ccgccttttt cccgagggtg 4861 ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg 4921 ccgccagaac acaggtaagt gccgtgtgtg gttcccgcgg gcctggcctc tttacgggtt 4981 atggcccttg cgtgccttga attacttcca cctggctgca gtacgtgatt cttgatcccg 5041 agcttcgggt tggaagtggg tgggagagtt cgaggccttg cgcttaagga gccccttcgc 5101 ctcgtgcttg agttgaggcc tggcctgggc gctggggccg ccgcgtgcga atctggtggc 5161 accttcgcgc ctgtctcgct gctttcgata agtctctagc catttaaaat ttttgatgac 5221 ctgctgcgac gctttttttc tggcaagata gtcttgtaaa tgcgggccaa gatctgcaca 5281 ctggtatttc ggtttttggg gccgcgggcg gcgacggggc ccgtgcgtcc cagcgcacat 5341 gttcggcgag gcggggcctg cgagcgcggc caccgagaat cggacggggg tagtctcaag 5401 ctggccggcc tgctctggtg cctggcctcg cgccgccgtg tatcgccccg ccctgggcgg 5461 caaggctggc ccggtcggca ccagttgcgt gagcggaaag atggccgctt cccggccctg 5521 ctgcagggag ctcaaaatgg aggacgcggc gctcgggaga gcgggcgggt gagtcaccca 5581 cacaaaggaa aagggccttt ccgtcctcag ccgtcgcttc atgtgactcc acggagtacc 5641 gggcgccgtc caggcacctc gattagttct cgagcttttg gagtacgtcg tctttaggtt 5701 ggggggaggg gttttatgcg atggagtttc cccacactga gtgggtggag actgaagtta 5761 ggccagcttg gcacttgatg taattctcct tggaatttgc cctttttgag tttggatctt 5821 ggttcattct caagcctcag acagtggttc aaagtttttt tcttccattt caggtgtcgt 5881 gaagcggccg ccatgctagt aaattgtccg ctaaattctg gccgtttttg gcttttttgt 5941 tagacggatc cataacttcg tataatgtat gctatacgaa gttatgccac catgtctaga 6001 ctggacaaga gcaaagtcat aaacggagct ctggaattac tcaatggtgt cggtatcgaa 6061 ggcctgacga caaggaaact cgctcaaaag ctgggagttg agcagcctac cctgtactgg 6121 cacgtgaaga acaagcgggc cctgctcgat gccctgccaa tcgagatgct ggacaggcat 6181 catacccact tctgccccct ggaaggcgag tcatggcaag actttctgcg gaacaacgcc 6241 aagtcatacc gctgtgctct cctctcacat cgcgacgggg ctaaagtgca tctcggcacc 6301 cgcccaacag agaaacagta cgaaaccctg gaaaatcagc tcgcgttcct gtgtcagcaa 6361 ggcttctccc tggagaacgc actgtacgct ctgtccgccg tgggccactt tacactgggc 6421 tgcgtattgg aggaacagga gcatcaagta gcaaaagagg aaagagagac acctaccacc 6481 gattctatgc ccccacttct gagacaagca attgagctgt tcgaccggca gggagccgaa 6541 cctgccttcc ttttcggcct ggaactaatc atatgtggcc tggagaaaca gctaaagtgc 6601 gaaagcggcg ggccgaccga cgcccttgac gattttgact tagacatgct cccagccgat 6661 gcccttgacg actttgacct tgatatgctg cctgctgacg ctcttgacga ttttgacctt 6721 gacatgctcc ccgggtaact aagtaaggat ccgcggccgc actagaggaa ttccgcccct 6781 ctcccccccc cccctctccc tccccccccc ctaacgttac tggccgaagc cgcttggaat 6841 aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct tttggcaatg 6901 tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt ctttcccctc 6961 tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct ctggaagctt 7021 cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc ccacctggcg 7081 acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag gcggcacaac 7141 cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc tcctcaagcg 7201 tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga tctgatctgg 7261 ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc 7321 cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataagcttac cggtccacca 7381 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg 7441 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg 7501 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc 7561 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc 7621 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct 7681 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg 7741 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca 7801 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg 7861 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg 7921 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact 7981 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc 8041 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaag 8101 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat 8161 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg 8221 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc 8281 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta 8341 tcgccttctt gacgagttct tctgatgtac aagtaaagcg gccgcgactc tagatcataa 8401 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc 8461 tgaacctgaa acataaaatg aatgcaattg ttgttgttta gtccctccca attcgatatc 8521 aagcttatcg cgatagatcc taatcaacct ctggattaca aaatttgtga aagattgact 8581 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg 8641 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg 8701 ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg 8761 tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct cctttccggg 8821 actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc 8881 tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca 8941 tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc 9001 tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct 9061 ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc 9121 gcctccccgc ctgagatcct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc 9181 actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc 9241 tgctttttgc ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg 9301 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag 9361 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag 9421 tgtggaaaat ctctagcagt agtagttcat gtcatcttat tattcagtat ttataacttg 9481 caaagaaatg aatatcagag agtgagaggc ccgggttaat taaggaaagg gctagatcat 9541 tcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata 9601 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt 9661 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg 9721 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt 9781 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta 9841 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc 9901 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa 9961 gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc 10021 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt 10081 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact 10141 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac 10201 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata 10261 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta 10321 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg 10381 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat 10441 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt 10501 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga 10561 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa 10621 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag 10681 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac 10741 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc 10801 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat 10861 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat 10921 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct 10981 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt 11041 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg 11101 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta 11161 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg 11221 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg 11281 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc 11341 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg 11401 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat 11461 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc 11521 agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg 11581 cgttggccga ttcattaatg cagcaagctc atggctgact aatttttttt atttatgcag 11641 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag 11701 gcctaggctt ttgcaaaaag ctccccgtgg cacgacaggt ttcccgactg gaaagcgggc 11761 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac 11821 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga 11881 aacagctatg acatgattac gaatttcaca aataaagcat ttttttcact gcattctagt 11941 tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatcaactg gataactcaa 12001 gctaaccaaa atcatcccaa acttcccacc ccatacccta ttaccactgc caattacctg 12061 tggtttcatt tactctaaac ctgtgattcc tctgaattat tttcatttta aagaaattgt 12121 atttgttaaa tatgtactac aaacttagta gt //
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