质粒名称: | pXR002: EF1a-dCasRx-2A-EGFP |
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出品公司: | 爱迪基因 |
目录编号: | 109050 |
存储实验室: | Patrick Hsu |
载体骨架基本信息 |
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载体名称: | lenti(AMP) |
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: | 12990 bp |
载体修饰: | -- |
载体类型: | 哺乳动物表达,慢病毒,CRISPR |
筛选标记: | GFP |
高拷贝/低拷贝: | 高拷贝 |
生长条件: | 37 ℃ |
插入基因信息 |
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插入基因名称: | dCasRx,NLS-dRfxCas13d-NLS |
物种来源: | H. sapiens (human); Ruminococcus flavefaciens XPD3002 |
插入大小: | 3000 bp |
基因突变: | R239A/H244A/R858A/H863A |
GenBank编号 | -- |
启动子 | -- |
标签/融合蛋白: |
NLS(插入件上的N端子)
NLS(插接件上的C端子)
HA(插入件上的C端子)
2A eGFP(主干上的C端)
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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: | pXR002: EF1a-dCasRx-2A-EGFP was a gift from Patrick Hsu (Addgene plasmid # 109050 ; http://n2t.net/addgene:109050 ; RRID:Addgene_109050) |
参考文献部分: | Transcriptome Engineering with RNA-Targeting Type VI-D CRISPR Effectors. Konermann S, Lotfy P, Brideau NJ, Oki J, Shokhirev MN, Hsu PD. Cell. 2018 Mar 8. pii: S0092-8674(18)30207-1. doi: 10.1016/j.cell.2018.02.033. 10.1016/j.cell.2018.02.033 PubMed 29551272 |
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pXR002: EF1a-dCasRx-2A-EGFP载体质粒基本信息
pXR002: EF1a-dCasRx-2A-EGFP质粒图谱载体图谱和pXR002: EF1a-dCasRx-2A-EGFP载体序列质粒序列多克隆位点信息
pXR002: EF1a-dCasRx-2A-EGFP质粒载体简介
pXR002: EF1a-dCasRx-2A-EGFP质粒序列载体序列
LOCUS pXR002: EF1a-dCasRx-2A-EGFP 12991 bp ds-DNA circular SYN 20-JUN-2019 DEFINITION Catalytically inactive dCasRx with 2A-EGFP. ACCESSION . VERSION . KEYWORDS pXR002: EF1a-dCasRx-2A-EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 12991) AUTHORS Konermann S, Lotfy P, Brideau NJ, Oki J, Shokhirev MN, Hsu PD TITLE Transcriptome Engineering with RNA-Targeting Type VI-D CRISPR Effectors. JOURNAL Cell. 2018 Mar 8. pii: S0092-8674(18)30207-1. doi: 10.1016/j.cell.2018.02.033. PUBMED 29551272 REFERENCE 2 (bases 1 to 12991) AUTHORS . TITLE Direct Submission JOURNAL Exported Jun 20, 2019 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..12991 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(42..61) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 233..612 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 614..812 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 767..787 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 830..1010 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 1057..1182 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1675..1908 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." misc_feature 2435..2552 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2677..3855 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 2908..3846 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 3803..3823 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" CDS 3874..3894 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 6811..6831 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 6841..6867 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 6883..6936 /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="EGRGSLLTCGDVEENPGP" CDS 6937..7653 /codon_start=1 /product="enhanced GFP" /label=EGFP /note="mammalian codon-optimized" /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" primer_bind complement(6979..7000) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(7240..7259) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 7587..7608 /label=EGFP-C /note="EGFP, forward primer" misc_feature 7678..8266 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(7731..7751) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(8149..8160) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(8269..8285) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(8270..8286) /label=pBluescriptKS /note="For pBluescript vector" LTR 8791..8971 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" primer_bind complement(8997..9014) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 9003..9227 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 9273..9701 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9360..9379) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 9570..9591 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(9710..9730) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 9715..10044 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 9895..10030 /label=SV40 ori /note="SV40 origin of replication" primer_bind 9957..9976 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 10092..10139 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 10158..10532 /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 10662..10783 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(10699..10718) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 10753..10772 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(10832..10848) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(10832..10848) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(10845..10867) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 10856..10872 /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(10880..10910) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 10925..10946 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(11063..11080) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(11234..11822) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(11314..11333) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(11993..12853) /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 12616..12635 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(12854..12958) /gene="bla" /label=AmpR promoter ORIGIN 1 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc 61 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg 121 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt 181 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga 241 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg 301 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc 361 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat 421 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat 481 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat 541 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc 601 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac 661 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa 721 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt 781 aggcgtgtac ggtgggaggt ctatataagc agcgcgtttt gcctgtactg ggtctctctg 841 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc 901 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg 961 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg 1021 aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt 1081 gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg 1141 actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga 1201 attagatcgc gatgggaaaa aattcggtta aggccagggg gaaagaaaaa atataaatta 1261 aaacatatag tatgggcaag cagggagcta gaacgattcg cagttaatcc tggcctgtta 1321 gaaacatcag aaggctgtag acaaatactg ggacagctac aaccatccct tcagacagga 1381 tcagaagaac ttagatcatt atataataca gtagcaaccc tctattgtgt gcatcaaagg 1441 atagagataa aagacaccaa ggaagcttta gacaagatag aggaagagca aaacaaaagt 1501 aagaccaccg cacagcaagc ggccgctgat cttcagacct ggaggaggag atatgaggga 1561 caattggaga agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc 1621 acccaccaag gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc 1681 tttgttcctt gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct 1741 gacggtacag gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag 1801 ggctattgag gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca 1861 ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg 1921 ttgctctgga aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa 1981 atctctggaa cagatttgga atcacacgac ctggatggag tgggacagag aaattaacaa 2041 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga 2101 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa 2161 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat 2221 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt 2281 tcagacccac ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg 2341 tggagagaga gacagagaca gatccattcg attagtgaac ggatcggcac tgcgtgcgcc 2401 aattctgcag acaaatggca gtattcatcc acaattttaa aagaaaaggg gggattgggg 2461 ggtacagtgc aggggaaaga atagtagaca taatagcaac agacatacaa actaaagaat 2521 tacaaaaaca aattacaaaa attcaaaatt ttcgggttta ttacagggac agcagagatc 2581 cagtttggtt aattaatgca aagatggata aagttttaaa cagagaggaa tctttgcagc 2641 taatggacct tctaggtctt gaaaggagtg ggaattggct ccggtgcccg tcagtgggca 2701 gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa ttgaaccggt 2761 gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt 2821 tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa cgttcttttt 2881 cgcaacgggt ttgccgccag aacacaggta agtgccgtgt gtggttcccg cgggcctggc 2941 ctctttacgg gttatggccc ttgcgtgcct tgaattactt ccacctggct gcagtacgtg 3001 attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc ttgcgcttaa 3061 ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg ccgccgcgtg 3121 cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct agccatttaa 3181 aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt aaatgcgggc 3241 caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg ggcccgtgcg 3301 tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag aatcggacgg 3361 gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc gtgtatcgcc 3421 ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga aagatggccg 3481 cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg agagcgggcg 3541 ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc ttcatgtgac 3601 tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgagctt ttggagtacg 3661 tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac tgagtgggtg 3721 gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt tgcccttttt 3781 gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt ttttcttcca 3841 tttcaggtgt cgtgacgtac ggccaccatg agccccaaga agaagagaaa ggtggaggcc 3901 agcatcgaaa aaaaaaagtc cttcgccaag ggcatgggcg tgaagtccac actcgtgtcc 3961 ggctccaaag tgtacatgac aaccttcgcc gaaggcagcg acgccaggct ggaaaagatc 4021 gtggagggcg acagcatcag gagcgtgaat gagggcgagg ccttcagcgc tgaaatggcc 4081 gataaaaacg ccggctataa gatcggcaac gccaaattca gccatcctaa gggctacgcc 4141 gtggtggcta acaaccctct gtatacagga cccgtccagc aggatatgct cggcctgaag 4201 gaaactctgg aaaagaggta cttcggcgag agcgctgatg gcaatgacaa tatttgtatc 4261 caggtgatcc ataacatcct ggacattgaa aaaatcctcg ccgaatacat taccaacgcc 4321 gcctacgccg tcaacaatat ctccggcctg gataaggaca ttattggatt cggcaagttc 4381 tccacagtgt atacctacga cgaattcaaa gaccccgagc accatagggc cgctttcaac 4441 aataacgata agctcatcaa cgccatcaag gcccagtatg acgagttcga caacttcctc 4501 gataacccca gactcggcta tttcggccag gcctttttca gcaaggaggg cagaaattac 4561 atcatcaatt acggcaacga atgctatgac attctggccc tcctgagcgg actggcgcac 4621 tgggtggtcg ctaacaacga agaagagtcc aggatctcca ggacctggct ctacaacctc 4681 gataagaacc tcgacaacga atacatctcc accctcaact acctctacga caggatcacc 4741 aatgagctga ccaactcctt ctccaagaac tccgccgcca acgtgaacta tattgccgaa 4801 actctgggaa tcaaccctgc cgaattcgcc gaacaatatt tcagattcag cattatgaaa 4861 gagcagaaaa acctcggatt caatatcacc aagctcaggg aagtgatgct ggacaggaag 4921 gatatgtccg agatcaggaa aaatcataag gtgttcgact ccatcaggac caaggtctac 4981 accatgatgg actttgtgat ttataggtat tacatcgaag aggatgccaa ggtggctgcc 5041 gccaataagt ccctccccga taatgagaag tccctgagcg agaaggatat ctttgtgatt 5101 aacctgaggg gctccttcaa cgacgaccag aaggatgccc tctactacga tgaagctaat 5161 agaatttgga gaaagctcga aaatatcatg cacaacatca aggaatttag gggaaacaag 5221 acaagagagt ataagaagaa ggacgcccct agactgccca gaatcctgcc cgctggccgt 5281 gatgtttccg ccttcagcaa actcatgtat gccctgacca tgttcctgga tggcaaggag 5341 atcaacgacc tcctgaccac cctgattaat aaattcgata acatccagag cttcctgaag 5401 gtgatgcctc tcatcggagt caacgctaag ttcgtggagg aatacgcctt tttcaaagac 5461 tccgccaaga tcgccgatga gctgaggctg atcaagtcct tcgctagaat gggagaacct 5521 attgccgatg ccaggagggc catgtatatc gacgccatcc gtattttagg aaccaacctg 5581 tcctatgatg agctcaaggc cctcgccgac accttttccc tggacgagaa cggaaacaag 5641 ctcaagaaag gcaagcacgg catgagaaat ttcattatta ataacgtgat cagcaataaa 5701 aggttccact acctgatcag atacggtgat cctgcccacc tccatgagat cgccaaaaac 5761 gaggccgtgg tgaagttcgt gctcggcagg atcgctgaca tccagaaaaa acagggccag 5821 aacggcaaga accagatcga caggtactac gaaacttgta tcggaaagga taagggcaag 5881 agcgtgagcg aaaaggtgga cgctctcaca aagatcatca ccggaatgaa ctacgaccaa 5941 ttcgacaaga aaaggagcgt cattgaggac accggcaggg aaaacgccga gagggagaag 6001 tttaaaaaga tcatcagcct gtacctcacc gtgatctacc acatcctcaa gaatattgtc 6061 aatatcaacg ccaggtacgt catcggattc cattgcgtcg agcgtgatgc tcaactgtac 6121 aaggagaaag gctacgacat caatctcaag aaactggaag agaagggatt cagctccgtc 6181 accaagctct gcgctggcat tgatgaaact gcccccgata agagaaagga cgtggaaaag 6241 gagatggctg aaagagccaa ggagagcatt gacagcctcg agagcgccaa ccccaagctg 6301 tatgccaatt acatcaaata cagcgacgag aagaaagccg aggagttcac caggcagatt 6361 aacagggaga aggccaaaac cgccctgaac gcctacctga ggaacaccaa gtggaatgtg 6421 atcatcaggg aggacctcct gagaattgac aacaagacat gtaccctgtt cgcaaacaag 6481 gccgtcgccc tggaagtggc caggtatgtc cacgcctata tcaacgacat tgccgaggtc 6541 aattcctact tccaactgta ccattacatc atgcagagaa ttatcatgaa tgagaggtac 6601 gagaaaagca gcggaaaggt gtccgagtac ttcgacgctg tgaatgacga gaagaagtac 6661 aacgataggc tcctgaaact gctgtgtgtg cctttcggct actgtatccc caggtttaag 6721 aacctgagca tcgaggccct gttcgatagg aacgaggccg ccaagttcga caaggagaaa 6781 aagaaggtgt ccggcaattc cggatccgga cctaagaaaa agaggaaggt ggcggccgct 6841 tacccatacg atgttccaga ttacgctgct agcggcagtg gagagggcag aggaagtctg 6901 ctaacatgcg gtgacgtcga ggagaatcct ggcccagtga gcaagggcga ggagctgttc 6961 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc 7021 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc 7081 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg 7141 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg 7201 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc 7261 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc 7321 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac 7381 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc 7441 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc 7501 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc 7561 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg 7621 atcactctcg gcatggacga gctgtacaag taagaattcg atatcaagct tatcgataat 7681 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct 7741 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg 7801 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg 7861 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt 7921 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt 7981 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg 8041 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc 8101 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat 8161 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc 8221 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga 8281 cctcgagacc tagaaaaaca tggagcaatc acaagtagca atacagcagc taccaatgct 8341 gattgtgcct ggctagaagc acaagaggag gaggaggtgg gttttccagt cacacctcag 8401 gtacctttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa 8461 aaggggggac tggaagggct aattcactcc caacgaagac aagatatcct tgatctgtgg 8521 atctaccaca cacaaggcta cttccctgat tggcagaact acacaccagg gccagggatc 8581 agatatccac tgacctttgg atggtgctac aagctagtac cagttgagca agagaaggta 8641 gaagaagcca atgaaggaga gaacacccgc ttgttacacc ctgtgagcct gcatgggatg 8701 gatgacccgg agagagaagt attagagtgg aggtttgaca gccgcctagc atttcatcac 8761 atggcccgag agctgcatcc ggactgtact gggtctctct ggttagacca gatctgagcc 8821 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga 8881 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga 8941 cccttttagt cagtgtggaa aatctctagc agggcccgtt taaacccgct gatcagcctc 9001 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac 9061 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg 9121 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga 9181 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga 9241 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc 9301 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc 9361 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct 9421 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa 9481 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc 9541 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact 9601 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg 9661 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt 9721 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat 9781 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg 9841 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg 9901 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt 9961 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt 10021 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct 10081 gatcagcacg tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca 10141 aggtgaggaa ctaaaccatg gccaagttga ccagtgccgt tccggtgctc accgcgcgcg 10201 acgtcgccgg agcggtcgag ttctggaccg accggctcgg gttctcccgg gacttcgtgg 10261 aggacgactt cgccggtgtg gtccgggacg acgtgaccct gttcatcagc gcggtccagg 10321 accaggtggt gccggacaac accctggcct gggtgtgggt gcgcggcctg gacgagctgt 10381 acgccgagtg gtcggaggtc gtgtccacga acttccggga cgcctccggg ccggccatga 10441 ccgagatcgg cgagcagccg tgggggcggg agttcgccct gcgcgacccg gccggcaact 10501 gcgtgcactt cgtggccgag gagcaggact gacacgtgct acgagatttc gattccaccg 10561 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc 10621 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt 10681 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac 10741 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt 10801 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt 10861 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg 10921 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg 10981 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc 11041 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc 11101 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata 11161 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg 11221 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct 11281 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa 11341 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 11401 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 11461 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 11521 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 11581 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 11641 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc 11701 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 11761 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 11821 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 11881 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa 11941 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat 12001 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct 12061 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg 12121 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag 12181 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta 12241 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg 12301 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg 12361 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct 12421 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta 12481 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg 12541 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc 12601 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg 12661 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga 12721 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg 12781 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat 12841 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc 12901 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca 12961 catttccccg aaaagtgcca cctgacgtcg a //