| 质粒名称: | pHAGE-TO-dCas9-3XGFP |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 64107 |
| 存储实验室: | Thoru Pederson |
|
载体骨架基本信息 |
|
| 载体名称: | pHAGE-DEST |
| 载体出品公司: | Clontech |
| 空载体大小: | 6000bp |
| 完整质粒大小: | 12980bp |
| 载体类型: | 哺乳细胞表达, Lentiviral |
| 筛选标记: | GFP |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | Stbl3 |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | Sp dCas9 |
| 物种来源: | Synthetic |
| 插入大小: | 4101bp |
| 启动子: | CMV-TO |
| 标签/融合蛋白: | 3XsfGFP (C terminal on insert) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | EcoRI (not destroyed) |
| 3’克隆位点: | XbaI (not destroyed) |
| 5’测序引物 | cgaaggaatagaagaagaaggtgga |
| 3’测序引物 | CCAAAGGGAGATCCGACTCG |
|
文献引用方法 |
|
| 材料和方法部分: |
pHAGE-TO-dCas9-3XGFP was a gift from Thoru Pederson (Addgene plasmid # 64107 ; http://n2t.net/addgene:64107 ; RRID:Addgene_64107)
|
| 参考文献部分: | Multicolor CRISPR labeling of chromosomal loci in human cells. Ma H, Naseri A, Reyes-Gutierrez P, Wolfe SA, Zhang S, Pederson T. Proc Natl Acad Sci U S A. 2015 Mar 10;112(10):3002-7. doi: 10.1073/pnas.1420024112. Epub 2015 Feb 23. 10.1073/pnas.1420024112 PubMed 25713381 |
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 pHAGE-TO-dCas9-3XGFP 12986 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Expression of Sp dCas9-3XGFP in mammalian cells.
ACCESSION .
VERSION .
KEYWORDS pHAGE-TO-dCas9-3XGFP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 12986)
AUTHORS Ma H, Naseri A, Reyes-Gutierrez P, Wolfe SA, Zhang S, Pederson T
TITLE Multicolor CRISPR labeling of chromosomal loci in human cells.
JOURNAL Proc Natl Acad Sci U S A. 2015 Mar 10;112(10):3002-7. doi:
10.1073/pnas.1420024112. Epub 2015 Feb 23.
PUBMED 25713381
REFERENCE 2 (bases 1 to 12986)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..12986
/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"
enhancer 2299..2602
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 2603..2806
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 2757..2777
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
protein_bind 2808..2826
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 2829..2847
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
primer_bind 2857..2881
/label=LNCX
/note="Human CMV promoter, forward primer"
regulatory 2974..2983
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2986..3006
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 3025..7125
/codon_start=1
/product="catalytically dead mutant of the Cas9
endonuclease from the Streptococcus pyogenes Type II
CRISPR/Cas system"
/label=dCas9
/note="RNA-guided DNA-binding protein that lacks
endonuclease activity due to the D10A mutation in the RuvC
catalytic domain and the H840A mutation in the HNH
catalytic domain"
/translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 7144..7164
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 7177..7890
/codon_start=1
/product="GFP variant that folds robustly even when fused
to poorly folded proteins (Pedelacq et al., 2006)"
/label=superfolder GFP
/translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
FVTAAGITHGMDELYK"
CDS 7909..8622
/codon_start=1
/product="GFP variant that folds robustly even when fused
to poorly folded proteins (Pedelacq et al., 2006)"
/label=superfolder GFP
/translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
FVTAAGITHGMDELYK"
CDS 8641..9354
/codon_start=1
/product="GFP variant that folds robustly even when fused
to poorly folded proteins (Pedelacq et al., 2006)"
/label=superfolder GFP
/translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
FVTAAGITHGMDELYK"
misc_feature 9376..9964
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(9429..9449)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(9847..9858)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
LTR 10039..10272
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
primer_bind complement(10391..10409)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 10477..10581
/gene="bla"
/label=AmpR promoter
CDS 10582..11442
/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(10800..10819)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 11613..12201
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 12102..12121
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 12355..12372
/label=L4440
/note="L4440 vector, forward primer"
primer_bind 12464..12483
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
protein_bind 12612..12633
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 12648..12678
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 12686..12702
/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 12691..12713
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 12710..12726
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 12710..12726
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind 12776..12795
/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 ttgacaaatg
2041 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
2101 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
2161 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gactagcgat
2221 gctagctgat gcgggcccta gttattaata gtaatcaatt acggggtcat tagttcatag
2281 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
2341 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
2401 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
2461 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
2521 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
2581 attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata
2641 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
2701 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
2761 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctcc ctatcagtga
2821 tagagatctc cctatcagtg atagagatcg tcgacgagct cgtttagtga accgtcagat
2881 cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg accgatccag
2941 cctaccggtt aaggtgaacg cgtgcaggct ggcgccacca tggcccctaa aaagaaacgc
3001 aaggtcgaat tcggaagcgg aagcgacaag aagtactcca ttgggctcgc tatcggtacc
3061 aacagcgtcg gctgggccgt cattacggac gagtacaagg tgccgagcaa aaaattcaaa
3121 gttctgggca ataccgatcg ccacagcata aagaagaacc tcattggagc cctcctgttc
3181 gactccgggg agacggccga agccacgcgg ctcaaaagaa cagcacggcg cagatatacc
3241 cgcagaaaga atcggatctg ctacctgcag gagatcttta gtaatgagat ggctaaggtg
3301 gatgactctt tcttccatag gctggaggag tcctttttgg tggaggagga taaaaagcac
3361 gagcgccacc caatctttgg caatatcgtg gacgaggtgg cgtaccatga aaagtaccca
3421 accatatatc atctgaggaa gaagctggta gacagtactg ataaggctga cttgcggttg
3481 atctatctcg cgctggcgca catgatcaaa tttcggggac acttcctcat cgagggggac
3541 ctgaacccag acaacagcga tgtcgacaaa ctctttatcc aactggttca gacttacaat
3601 cagcttttcg aggagaaccc gatcaacgca tccggcgttg acgccaaagc aatcctgagc
3661 gctaggctgt ccaaatcccg gcggctcgaa aacctcatcg cacagctccc tggggagaag
3721 aagaacggcc tgtttggtaa tcttatcgcc ctgtcactcg ggctgacccc caactttaaa
3781 tctaacttcg acctggccga agatgccaag ctgcaactga gcaaagacac ctacgatgat
3841 gatctcgaca atctgctggc ccagatcggc gaccagtacg cagacctttt tttggcggca
3901 aagaacctgt cagacgccat tctgctgagt gatattctgc gagtgaacac ggagatcacc
3961 aaagctccgc tgagcgctag tatgatcaag cgctatgatg agcaccacca agacttgact
4021 ttgctgaagg cccttgtcag acagcaactg cctgagaagt acaaggaaat tttcttcgat
4081 cagtctaaaa atggctacgc cggatacatt gacggcggag caagccagga ggaattttac
4141 aaatttatta agcccatctt ggaaaaaatg gacggcaccg aggagctgct ggtaaagctg
4201 aacagagaag atctgttgcg caaacagcgc actttcgaca atggaagcat cccccaccag
4261 attcacctgg gcgaactgca cgctatcctc aggcggcaag aggatttcta cccctttttg
4321 aaagataaca gggaaaagat tgagaaaatc ctcacatttc ggatacccta ctatgtaggc
4381 cccctcgctc ggggaaattc cagattcgcg tggatgactc gcaaatcaga agagaccatc
4441 actccctgga acttcgagga agtcgtggat aagggggcct ctgcccagtc cttcatcgaa
4501 aggatgacta actttgataa aaatctgcct aacgaaaagg tgcttcctaa acactctctg
4561 ctgtacgagt acttcacagt ttataacgag ctcaccaagg tcaaatacgt cacagaaggg
4621 atgagaaagc cagcattcct gtctggagag cagaagaaag ctatcgtgga cctcctcttc
4681 aagacgaacc ggaaagttac cgtgaaacag ctcaaagaag actatttcaa aaagattgaa
4741 tgtttcgact ctgttgaaat cagcggagtg gaggatcgct tcaacgcatc cctgggaacg
4801 tatcacgatc tcctgaaaat cattaaagac aaggacttcc tggacaatga ggagaacgag
4861 gacattcttg aggacattgt cctcaccctt acgttgtttg aagataggga gatgattgaa
4921 gaacgcttga aaacttacgc tcatctcttc gacgacaaag tcatgaaaca gctcaagaga
4981 cgccgatata caggatgggg gcggctgtca agaaaactga tcaatggcat ccgagacaag
5041 cagagtggaa agacaatcct ggattttctt aagtccgatg gatttgccaa ccggaacttc
5101 atgcagttga tccatgatga ctctctcacc tttaaggagg acatccagaa agcacaagtt
5161 tctggccagg gggacagtct tcacgagcac atcgctaatc ttgcaggtag cccagctatc
5221 aaaaagggaa tactgcagac cgttaaggtc gtggatgaac tcgtcaaagt aatgggaagg
5281 cataagcccg agaatatcgt tatcgagatg gcccgagaga accaaactac ccagaaggga
5341 cagaagaaca gtagggaaag gatgaagagg attgaagagg gtataaaaga actggggtcc
5401 caaatcctta aggaacaccc agttgaaaac acccagcttc agaatgagaa gctctacctg
5461 tactacctgc agaacggcag ggacatgtac gtggatcagg aactggacat caaccggttg
5521 tccgactacg acgtggatgc tatcgtgccc caaagctttc tcaaagatga ttctattgat
5581 aataaagtgt tgacaagatc cgataaaaat agagggaaga gtgataacgt cccctcagaa
5641 gaagttgtca agaaaatgaa aaattattgg cggcagctgc tgaacgccaa actgatcaca
5701 caacggaagt tcgataatct gactaaggct gaacgaggtg gcctgtctga gttggataaa
5761 gccggcttca tcaaaaggca gcttgttgag acacgccaga tcaccaagca cgtggcccaa
5821 attctcgatt cacgcatgaa caccaagtac gatgaaaatg acaaactgat tcgagaggtg
5881 aaagttatta ctctgaagtc taagctggtc tcagatttca gaaaggactt tcagttttat
5941 aaggtgagag agatcaacaa ttaccaccat gcgcatgatg cctacctgaa tgcagtggta
6001 ggcactgcac ttatcaaaaa atatcccaag ctggaatctg aatttgttta cggagactat
6061 aaagtgtacg atgttaggaa aatgatcgca aagtctgagc aggaaatagg caaggccacc
6121 gctaagtact tcttttacag caatattatg aattttttca agaccgagat tacactggcc
6181 aatggagaga ttcggaagcg accacttatc gaaacaaacg gagaaacagg agaaatcgtg
6241 tgggacaagg gtagggattt cgcgacagtc cgcaaggtcc tgtccatgcc gcaggtgaac
6301 atcgttaaaa agaccgaagt acagaccgga ggcttctcca aggaaagtat cctcccgaaa
6361 aggaacagcg acaagctgat cgcacgcaaa aaagattggg accccaagaa atacggcgga
6421 ttcgattctc ctacagtcgc ttacagtgta ctggttgtgg ccaaagtgga gaaagggaag
6481 tctaaaaaac tcaaaagcgt caaggaactg ctgggcatca caatcatgga gcgatccagc
6541 ttcgagaaaa accccatcga ctttctcgaa gcgaaaggat ataaagaggt caaaaaagac
6601 ctcatcatta agctgcccaa gtactctctc tttgagcttg aaaacggccg gaaacgaatg
6661 ctcgctagtg cgggcgagct gcagaaaggt aacgagctgg cactgccctc taaatacgtt
6721 aatttcttgt atctggccag ccactatgaa aagctcaaag ggtctcccga agataatgag
6781 cagaagcagc tgttcgtgga acaacacaaa cactaccttg atgagatcat cgagcaaata
6841 agcgagttct ccaaaagagt gatcctcgcc gacgctaacc tcgataaggt gctttctgct
6901 tacaataagc acagggataa gcccatcagg gagcaggcag aaaacattat ccacttgttt
6961 actctgacca acttgggcgc gcctgcagcc ttcaagtact tcgacaccac catagacaga
7021 aagcggtaca cctctacaaa ggaggtcctg gacgccacac tgattcatca gtcaattacg
7081 gggctctatg aaacaagaat cgacctctct cagctcggtg gagacggctc tactagtggc
7141 tctcccaaga agaagaggaa ggtaggatcc ggaagtatgc gtaaaggcga agagctgttc
7201 actggtgtcg tccctattct ggtggaactg gatggtgatg tcaacggtca taagttttcc
7261 gtgcgtggcg agggtgaagg tgacgcaact aatggtaaac tgacgctgaa gttcatctgt
7321 actactggta aactgccggt accttggccg actctggtaa cgacgctgac ttatggtgtt
7381 cagtgctttg ctcgttatcc ggaccatatg aagcagcatg acttcttcaa gtccgccatg
7441 ccggaaggct atgtgcagga acgcacgatt tcctttaagg atgacggcac gtacaaaacg
7501 cgtgcggaag tgaaatttga aggcgatacc ctggtaaacc gcattgagct gaaaggcatt
7561 gactttaaag aagacggcaa tatcctgggc cataagctgg aatacaattt taacagccac
7621 aatgtttaca tcaccgccga taaacaaaaa aatggcatta aagcgaattt taaaattcgc
7681 cacaacgtgg aggatggcag cgtgcagctg gctgatcact accagcaaaa cactccaatc
7741 ggtgatggtc ctgttctgct gccagacaat cactatctga gcacgcaaag cgttctgtct
7801 aaagatccga acgagaaacg cgatcatatg gttctgctgg agttcgtaac cgcagcgggc
7861 atcacgcatg gtatggatga actgtacaaa tctggcggcc gctctggcat gcgtaaaggc
7921 gaagagctgt tcactggtgt cgtccctatt ctggtggaac tggatggtga tgtcaacggt
7981 cataagtttt ccgtgcgtgg cgagggtgaa ggtgacgcaa ctaatggtaa actgacgctg
8041 aagttcatct gtactactgg taaactgccg gtaccttggc cgactctggt aacgacgctg
8101 acttatggtg ttcagtgctt tgctcgttat ccggaccata tgaagcagca tgacttcttc
8161 aagtccgcca tgccggaagg ctatgtgcag gaacgcacga tttcctttaa ggatgacggc
8221 acgtacaaaa cgcgtgcgga agtgaaattt gaaggcgata ccctggtaaa ccgcattgag
8281 ctgaaaggca ttgactttaa agaagacggc aatatcctgg gccataagct ggaatacaat
8341 tttaacagcc acaatgttta catcaccgcc gataaacaaa aaaatggcat taaagcgaat
8401 tttaaaattc gccacaacgt ggaggatggc agcgtgcagc tggctgatca ctaccagcaa
8461 aacactccaa tcggtgatgg tcctgttctg ctgccagaca atcactatct gagcacgcaa
8521 agcgttctgt ctaaagatcc gaacgagaaa cgcgatcata tggttctgct ggagttcgta
8581 accgcagcgg gcatcacgca tggtatggat gaactgtaca aaggtagcat cgatggtagc
8641 atgcgtaaag gcgaagagct gttcactggt gtcgtcccta ttctggtgga actggatggt
8701 gatgtcaacg gtcataagtt ttccgtgcgt ggcgagggtg aaggtgacgc aactaatggt
8761 aaactgacgc tgaagttcat ctgtactact ggtaaactgc cggtaccttg gccgactctg
8821 gtaacgacgc tgacttatgg tgttcagtgc tttgctcgtt atccggacca tatgaagcag
8881 catgacttct tcaagtccgc catgccggaa ggctatgtgc aggaacgcac gatttccttt
8941 aaggatgacg gcacgtacaa aacgcgtgcg gaagtgaaat ttgaaggcga taccctggta
9001 aaccgcattg agctgaaagg cattgacttt aaagaagacg gcaatatcct gggccataag
9061 ctggaataca attttaacag ccacaatgtt tacatcaccg ccgataaaca aaaaaatggc
9121 attaaagcga attttaaaat tcgccacaac gtggaggatg gcagcgtgca gctggctgat
9181 cactaccagc aaaacactcc aatcggtgat ggtcctgttc tgctgccaga caatcactat
9241 ctgagcacgc aaagcgttct gtctaaagat ccgaacgaga aacgcgatca tatggttctg
9301 ctggagttcg taaccgcagc gggcatcacg catggtatgg atgaactgta caaactcgag
9361 tagggtaatc tagataatca acctctggat tacaaaattt gtgaaagatt gactggtatt
9421 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
9481 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
9541 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
9601 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
9661 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
9721 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
9781 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
9841 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
9901 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
9961 ccgcctgaga tcctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
10021 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa gatctgcttt
10081 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
10141 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
10201 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
10261 aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga
10321 aatgaatatc agagagtgag aggcccgggt taattaagga aagggctaga tcattcttga
10381 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
10441 tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
10501 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
10561 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
10621 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
10681 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
10741 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
10801 ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata
10861 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
10921 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
10981 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
11041 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
11101 gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact
11161 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
11221 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
11281 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
11341 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
11401 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
11461 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
11521 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
11581 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
11641 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
11701 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
11761 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
11821 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
11881 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
11941 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
12001 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
12061 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
12121 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
12181 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
12241 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
12301 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
12361 tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg
12421 ccgattcatt aatgcagcaa gctcatggct gactaatttt ttttatttat gcagaggccg
12481 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
12541 gcttttgcaa aaagctcccc gtggcacgac aggtttcccg actggaaagc gggcagtgag
12601 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
12661 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
12721 tatgacatga ttacgaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
12781 ttgtccaaac tcatcaatgt atcttatcat gtctggatca actggataac tcaagctaac
12841 caaaatcatc ccaaacttcc caccccatac cctattacca ctgccaatta cctgtggttt
12901 catttactct aaacctgtga ttcctctgaa ttattttcat tttaaagaaa ttgtatttgt
12961 taaatatgta ctacaaactt agtagt
//
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