质粒名称: | pSLIK-Venus |
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出品公司: | Iain Fraser |
目录编号: | 25734 |
存储实验室: | Iain Fraser |
载体骨架基本信息 |
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载体名称: | pSLIK |
载体出品公司: | Iain Fraser |
完整质粒大小: | 13170bp |
载体修饰: | DB3.1 (ccdB survival) |
载体类型: | 哺乳细胞表达 |
筛选标记: | Venus |
载体抗性: | Chloramphenicol and Ampicillin, 25 & 100 μg/mL |
生长温度: | 37 ℃ |
宿主菌: | ccdB Survival |
拷贝数: | 未知 |
插入基因信息 |
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克隆方法: | Gateway Cloning |
5’克隆位点: | attR1(未破坏) |
3’克隆位点: | attR2 (未破坏) |
5’测序引物 | CMV-F |
3’测序引物 | EXFP-R |
文献引用方法 |
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材料和方法部分: | pSLIK-Venus was a gift from Iain Fraser (Addgene plasmid # 25734 ; http://n2t.net/addgene:25734 ; RRID:Addgene_25734) |
参考文献部分: | A single lentiviral vector platform for microRNA-based conditional RNA interference and coordinated transgene expression. Shin KJ, Wall EA, Zavzavadjian JR, Santat LA, Liu J, Hwang JI, Rebres R, Roach T, Seaman W, Simon MI, Fraser ID. Proc Natl Acad Sci U S A. 2006 Sep 12. 103(37):13759-64. 10.1073/pnas.0606179103 PubMed 16945906 |
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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 pSLIK-Venus 13170 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Lentiviral vector for Tet-based inducible shRNA or cDNA expression, constitutive Venus fluorescent protein coexpression.. ACCESSION . VERSION . KEYWORDS pSLIK-Venus SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 13170) AUTHORS Shin KJ, Wall EA, Zavzavadjian JR, Santat LA, Liu J, Hwang JI, Rebres R, Roach T, Seaman W, Simon MI, Fraser ID TITLE A single lentiviral vector platform for microRNA-based conditional RNA interference and coordinated transgene expression. JOURNAL Proc Natl Acad Sci U S A. 2006 Sep 12. 103(37):13759-64. PUBMED 16945906 REFERENCE 2 (bases 1 to 13170) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.biofeng.com FEATURES Location/Qualifiers source 1..13170 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(47..66) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 238..617 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 619..817 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 772..792 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 835..1015 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 1062..1187 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1680..1913 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 2098..2142 /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 2440..2557 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" protein_bind 2614..2738 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 2763..2793 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 2847..3506 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind complement(2913..2932) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 3848..4153 /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 4032..4051 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(4194..4318) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 4368..5579 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 5416..5435 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 5704..6450 /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 6488..7062 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(6655..6672) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 6882..6901 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 7090..7809 /codon_start=1 /product="yellow fluorescent protein (YFP) with fast and efficient maturation (Nagai et al., 2002)" /label=Venus /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(7135..7156) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(7396..7415) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 7743..7764 /label=EGFP-C /note="EGFP, forward primer" misc_feature 7858..8446 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(7911..7931) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(8329..8340) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(8449..8465) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(8450..8466) /label=pBluescriptKS /note="For pBluescript vector" LTR 8975..9155 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" primer_bind complement(9181..9198) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 9187..9411 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 9457..9885 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9544..9563) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 9754..9775 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(9894..9914) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 9899..10228 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 10079..10214 /label=SV40 ori /note="SV40 origin of replication" primer_bind 10141..10160 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 10276..10323 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 10342..10716 /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 10846..10967 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(10883..10902) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 10937..10956 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(11016..11032) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(11016..11032) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(11029..11051) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 11040..11056 /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(11064..11094) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 11109..11130 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(11247..11264) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(11418..12006) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(11498..11517) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(12177..13037) /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 12800..12819 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(13038..13142) /gene="bla" /label=AmpR promoter ORIGIN 1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg 61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt 121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc 181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac 241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat 301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg 361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt 421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag 481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc 541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag 601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt 661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc 721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg 781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct 841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt 901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac 961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc 1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc 1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa 1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg 1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata 1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc 1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga 1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc 1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca 1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg 1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga 1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata 1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg 1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg 1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag 1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt 1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt 1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt 2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag 2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata 2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta 2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta 2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa 2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt 2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat 2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa 2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag 2581 agatccagtt tggttaattc tagacagtta tcaacaagtt tgtacaaaaa agctgaacga 2641 gaaacgtaaa atgatataaa tatcaatata ttaaattaga ttttgcataa aaaacagact 2701 acataatact gtaaaacaca acatatccag tcactatggc ggccgcatta ggcaccccag 2761 gctttacact ttatgcttcc ggctcgtata atgtgtggat tttgagttag gatccggcga 2821 gattttcagg agctaaggaa gctaaaatgg agaaaaaaat cactggatat accaccgttg 2881 atatatccca atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta 2941 cctataacca gaccgttcag ctggatatta cggccttttt aaagaccgta aagaaaaata 3001 agcacaagtt ttatccggcc tttattcaca ttcttgcccg cctgatgaat gctcatccgg 3061 aattccgtat ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt cacccttgtt 3121 acaccgtttt ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa taccacgacg 3181 atttccggca gtttctacac atatattcgc aagatgtggc gtgttacggt gaaaacctgg 3241 cctatttccc taaagggttt attgagaata tgtttttcgt ctcagccaat ccctgggtga 3301 gtttcaccag ttttgattta aacgtggcca atatggacaa cttcttcgcc cccgttttca 3361 ccatgggcaa atattatacg caaggcgaca aggtgctgat gccgctggcg attcaggttc 3421 atcatgccgt ctgtgatggc ttccatgtcg gcagaatgct taatgaatta caacagtact 3481 gcgatgagtg gcagggcggg gcgtaaagat ctggatccgg cttactaaaa gccagataac 3541 agtatgcgta tttgcgcgct gatttttgcg gtataagaat atatactgat atgtataccc 3601 gaagtatgtc aaaaagaggt gtgctatgaa gcagcgtatt acagtgacag ttgacagcga 3661 cagctatcag ttgctcaagg catatatgat gtcaatatct ccggtctggt aagcacaacc 3721 atgcagaatg aagcccgtcg tctgcgtgcc gaacgctgga aagcggaaaa tcaggaaggg 3781 atggctgagg tcgcccggtt tattgaaatg aacggctctt ttgctgacga gaacagggac 3841 tggtgaaatg cagtttaagg tttacaccta taaaagagag agccgttatc gtctgtttgt 3901 ggatgtacag agtgatatta ttgacacgcc cgggcgacgg atggtgatcc ccctggccag 3961 tgcacgtctg ctgtcagata aagtctcccg tgaactttac ccggtggtgc atatcgggga 4021 tgaaagctgg cgcatgatga ccaccgatat ggccagtgtg ccggtctccg ttatcgggga 4081 agaagtggct gatctcagcc accgcgaaaa tgacatcaaa aacgccatta acctgatgtt 4141 ctggggaata taaatgtcag gctcccttat acacagccag tctgcaggtc gaccatagtg 4201 actggatatg ttgtgtttta cagtattatg tagtctgttt tttatgcaaa atctaattta 4261 atatattgat atttatatca ttttacgttt ctcgttcagc tttcttgtac aaagtggttg 4321 ataacatctc gagccataaa gatggttaat taacccaccc aagatctggc ctccgcgccg 4381 ggttttggcg cctcccgcgg gcgcccccct cctcacggcg agcgctgcca cgtcagacga 4441 agggcgcagc gagcgtcctg atccttccgc ccggacgctc aggacagcgg cccgctgctc 4501 ataagactcg gccttagaac cccagtatca gcagaaggac attttaggac gggacttggg 4561 tgactctagg gcactggttt tctttccaga gagcggaaca ggcgaggaaa agtagtccct 4621 tctcggcgat tctgcggagg gatctccgtg gggcggtgaa cgccgatgat tatataagga 4681 cgcgccgggt gtggcacagc tagttccgtc gcagccggga tttgggtcgc ggttcttgtt 4741 tgtggatcgc tgtgatcgtc acttggtgag tagcgggctg ctgggctggc cggggctttc 4801 gtggccgccg ggccgctcgg tgggacggaa gcgtgtggag agaccgccaa gggctgtagt 4861 ctgggtccgc gagcaaggtt gccctgaact gggggttggg gggagcgcag caaaatggcg 4921 gctgttcccg agtcttgaat ggaagacgct tgtgaggcgg gctgtgaggt cgttgaaaca 4981 aggtgggggg catggtgggc ggcaagaacc caaggtcttg aggccttcgc taatgcggga 5041 aagctcttat tcgggtgaga tgggctgggg caccatctgg ggaccctgac gtgaagtttg 5101 tcactgactg gagaactcgg tttgtcgtct gttgcggggg cggcagttat ggcggtgccg 5161 ttgggcagtg cacccgtacc tttgggagcg cgcgccctcg tcgtgtcgtg acgtcacccg 5221 ttctgttggc ttataatgca gggtggggcc acctgccggt aggtgtgcgg taggcttttc 5281 tccgtcgcag gacgcagggt tcgggcctag ggtaggctct cctgaatcga caggcgccgg 5341 acctctggtg aggggaggga taagtgaggc gtcagtttct ttggtcggtt ttatgtacct 5401 atcttcttaa gtagctgaag ctccggtttt gaactatgcg ctcggggttg gcgagtgtgt 5461 tttgtgaagt tttttaggca ccttttgaaa tgtaatcatt tgggtcaata tgtaattttc 5521 agtgttagac tagtaaattg tccgctaaat tctggccgtt tttggctttt ttgttagacg 5581 aagcttggta ccgagctcgg atctccaccc cgtaccggtc ctgcaggatt taaatggcgc 5641 gccgctagcg ctaccggact cagatctcga gctcaagctt cgaattctgc agtcgaattc 5701 accatgtcta gactggacaa gagcaaagtc ataaacggag ctctggaatt actcaatggt 5761 gtcggtatcg aaggcctgac gacaaggaaa ctcgctcaaa agctgggagt tgagcagcct 5821 accctgtact ggcacgtgaa gaacaagcgg gccctgctcg atgccctgcc aatcgagatg 5881 ctggacaggc atcataccca cttctgcccc ctggaaggcg agtcatggca agactttctg 5941 cggaacaacg ccaagtcata ccgctgtgct ctcctctcac atcgcgacgg ggctaaagtg 6001 catctcggca cccgcccaac agagaaacag tacgaaaccc tggaaaatca gctcgcgttc 6061 ctgtgtcagc aaggcttctc cctggagaac gcactgtacg ctctgtccgc cgtgggccac 6121 tttacactgg gctgcgtatt ggaggaacag gagcatcaag tagcaaaaga ggaaagagag 6181 acacctacca ccgattctat gcccccactt ctgagacaag caattgagct gttcgaccgg 6241 cagggagccg aacctgcctt ccttttcggc ctggaactaa tcatatgtgg cctggagaaa 6301 cagctaaagt gcgaaagcgg cgggccgacc gacgcccttg acgattttga cttagacatg 6361 ctcccagccg atgcccttga cgactttgac cttgatatgc tgcctgctga cgctcttgac 6421 gattttgacc ttgacatgct ccccgggtaa ctaagtaagg atccgcggcc gcactagagg 6481 aattccgccc ctctccctcc ccccccccta acgttactgg ccgaagccgc ttggaataag 6541 gccggtgtgc gtttgtctat atgttatttt ccaccatatt gccgtctttt ggcaatgtga 6601 gggcccggaa acctggccct gtcttcttga cgagcattcc taggggtctt tcccctctcg 6661 ccaaaggaat gcaaggtctg ttgaatgtcg tgaaggaagc agttcctctg gaagcttctt 6721 gaagacaaac aacgtctgta gcgacccttt gcaggcagcg gaacccccca cctggcgaca 6781 ggtgcctctg cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc 6841 agtgccacgt tgtgagttgg atagttgtgg aaagagtcaa atggctctcc tcaagcgtat 6901 tcaacaaggg gctgaaggat gcccagaagg taccccattg tatgggattc tgatctgggg 6961 cctcggtgca catgctttac atgtgtttag tcgaggttaa aaaaacgtct aggccccccg 7021 aaccacgggg acgtggtttt cctttgaaaa acacgatgat aagcttaccg gtacgcgtga 7081 tgatccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 7141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat 7201 gccacctacg gcaagctgac cctgaagctg atctgcacca ccggcaagct gcccgtgccc 7261 tggcccaccc tcgtgaccac cctgggctac ggcctgcagt gcttcgcccg ctaccccgac 7321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 7381 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 7441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 7501 ctggggcaca agctggagta caactacaac agccacaacg tctatatcac cgccgacaag 7561 cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga cggcggcgtg 7621 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 7681 gacaaccact acctgagcta ccagtccgcc ctgagcaaag accccaacga gaagcgcgat 7741 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg 7801 tacaagtaag aattgttgtt gtttagtccc tcccaattcg atatcaagct tatcgataat 7861 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct 7921 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg 7981 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg 8041 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt 8101 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt 8161 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg 8221 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc 8281 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat 8341 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc 8401 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga 8461 cctcgatcga gacctagaaa aacatggagc aatcacaagt agcaatacag cagctaccaa 8521 tgctgattgt gcctggctag aagcacaaga ggaggaggag gtgggttttc cagtcacacc 8581 tcaggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa 8641 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagata tccttgatct 8701 gtggatctac cacacacaag gctacttccc tgattggcag aactacacac cagggccagg 8761 gatcagatat ccactgacct ttggatggtg ctacaagcta gtaccagttg agcaagagaa 8821 ggtagaagaa gccaatgaag gagagaacac ccgcttgtta caccctgtga gcctgcatgg 8881 gatggatgac ccggagagag aagtattaga gtggaggttt gacagccgcc tagcatttca 8941 tcacatggcc cgagagctgc atccggactg tactgggtct ctctggttag accagatctg 9001 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc 9061 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct 9121 cagacccttt tagtcagtgt ggaaaatctc tagcagggcc cgtttaaacc cgctgatcag 9181 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct 9241 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc 9301 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg 9361 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg 9421 cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa 9481 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc 9541 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag 9601 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca 9661 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc 9721 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa 9781 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct 9841 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt 9901 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat 9961 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag 10021 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat 10081 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt 10141 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg 10201 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg 10261 atctgatcag cacgtgttga caattaatca tcggcatagt atatcggcat agtataatac 10321 gacaaggtga ggaactaaac catggccaag ttgaccagtg ccgttccggt gctcaccgcg 10381 cgcgacgtcg ccggagcggt cgagttctgg accgaccggc tcgggttctc ccgggacttc 10441 gtggaggacg acttcgccgg tgtggtccgg gacgacgtga ccctgttcat cagcgcggtc 10501 caggaccagg tggtgccgga caacaccctg gcctgggtgt gggtgcgcgg cctggacgag 10561 ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc gggacgcctc cgggccggcc 10621 atgaccgaga tcggcgagca gccgtggggg cgggagttcg ccctgcgcga cccggccggc 10681 aactgcgtgc acttcgtggc cgaggagcag gactgacacg tgctacgaga tttcgattcc 10741 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg 10801 atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca 10861 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt 10921 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata 10981 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat 11041 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg 11101 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag 11161 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt 11221 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg 11281 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg 11341 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag 11401 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga 11461 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct 11521 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc 11581 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg 11641 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc 11701 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca 11761 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag 11821 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct 11881 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc 11941 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga 12001 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca 12061 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat 12121 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac 12181 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt 12241 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt 12301 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag 12361 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct 12421 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt 12481 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc 12541 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt 12601 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg 12661 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg 12721 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct 12781 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc 12841 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt 12901 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt 12961 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg 13021 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat 13081 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg 13141 cgcacatttc cccgaaaagt gccacctgac //
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