| 质粒名称: | pX459V2.0+K848A |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 108296 |
| 存储实验室: | Yuichiro Miyaoka |
|
载体骨架基本信息 |
|
| 载体名称: | pX459 |
| 载体出品公司: | Dr. Feng Zhang's Lab |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 9175 bp |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达, CRISPR |
| 筛选标记: | 嘌呤霉素 |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 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: | 限制性内切酶 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | ACTATCATATGCTTACCGTAAC |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pX459V2.0+K848A was a gift from Yuichiro Miyaoka (Addgene plasmid # 108296 ; http://n2t.net/addgene:108296 ; RRID:Addgene_108296) |
| 参考文献部分: | Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 with improved proof-reading enhances homology-directed repair. Tomoko Kato-Inui, Gou Takahashi, Szuyin Hsu, Yuichiro Miyaoka. Nucleic Acids Research 10.1093/nar/gky264 |
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 pX459V2.0+K848A 9174 bp ds-DNA circular SYN 07-AUG-2018
DEFINITION pX459 V2.0 (Plasmid #62988) with the K848A mutation.
ACCESSION .
VERSION .
KEYWORDS pX459V2.0+K848A
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9174)
AUTHORS Tomoko Kato-Inui, Gou Takahashi, Szuyin Hsu, Yuichiro Miyaoka
TITLE Clustered regularly interspaced short palindromic repeats
(CRISPR)/CRISPR-associated protein 9 with improved proof-reading
enhances homology-directed repair
JOURNAL Nucleic Acids Research
REFERENCE 2 (bases 1 to 9174)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 7, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..9174
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..241
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 1..21
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 172..191
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 268..343
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
enhancer 440..725
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer;
contains an 18-bp deletion relative to the standard CMV
enhancer"
promoter 727..1004
/label=chicken beta-actin promoter
intron 1005..1232
/label=hybrid intron
/note="hybrid between chicken beta-actin (CBA) and minute
virus of mice (MMV) introns (Gray et al., 2011)"
regulatory 1244..1253
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 1253..1318
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS 1325..1345
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 1370..5470
/codon_start=1
/product="Cas9 endonuclease from the Streptococcus pyogenes
Type II CRISPR/Cas system, mutated to improve targeting
specificity (Slaymaker et al., 2016)"
/label=eSpCas9(1.1)
/note="carries the mutations K848A, K1003A, and R1060A"
/translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLADDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 1370..5470
/codon_start=1
/product="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
/label=Cas9
/note="generates RNA-guided double strand breaks in DNA"
/translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLADDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 5471..5518
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS 5534..5587
/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 5588..6187
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind complement(5588..5607)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
primer_bind 6084..6104
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
primer_bind complement(6215..6232)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 6221..6428
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
repeat_region 6437..6577
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 6437..6566
/label=AAV2 ITR
rep_origin 6652..7107
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(6739..6758)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 6949..6970
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(7124..7143)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 7243..7265
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(7303..7321)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 7389..7493
/gene="bla"
/label=AmpR promoter
CDS 7494..8354
/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 complement(7712..7731)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 8525..9113
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 9014..9033
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
241 cgaaacaccg ggtcttcgag aagacctgtt ttagagctag aaatagcaag ttaaaataag
301 gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttg ttttagagct
361 agaaatagca agttaaaata aggctagtcc gtttttagcg cgtgcgccaa ttctgcagac
421 aaatggctct agaggtaccc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
481 ccaacgaccc ccgcccattg acgtcaatag taacgccaat agggactttc cattgacgtc
541 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
601 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tgtgcccagt
661 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
721 ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc ccctccccac
781 ccccaatttt gtatttattt attttttaat tattttgtgc agcgatgggg gcgggggggg
841 ggggggggcg cgcgccaggc gggggggggg ggggcggggg gggggggggg gcggggcgga
901 gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa gtttcctttt atggcgaggc
961 ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc gctgcgcgct
1021 gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg cccgccccgg ctctgactga
1081 ccgcgttact cccacaggtg agcgggcggg acggcccttc tcctccgggc tgtaattagc
1141 tgagcaagag gtaagggttt aagggatggt tggttggtgg ggtattaatg tttaattacc
1201 tggagcacct gcctgaaatc actttttttc aggttggacc ggtgccacca tggactataa
1261 ggaccacgac ggagactaca aggatcatga tattgattac aaagacgatg acgataagat
1321 ggccccaaag aagaagcgga aggtcggtat ccacggagtc ccagcagccg acaagaagta
1381 cagcatcggc ctggacatcg gcaccaactc tgtgggctgg gccgtgatca ccgacgagta
1441 caaggtgccc agcaagaaat tcaaggtgct gggcaacacc gaccggcaca gcatcaagaa
1501 gaacctgatc ggagccctgc tgttcgacag cggcgaaaca gccgaggcca cccggctgaa
1561 gagaaccgcc agaagaagat acaccagacg gaagaaccgg atctgctatc tgcaagagat
1621 cttcagcaac gagatggcca aggtggacga cagcttcttc cacagactgg aagagtcctt
1681 cctggtggaa gaggataaga agcacgagcg gcaccccatc ttcggcaaca tcgtggacga
1741 ggtggcctac cacgagaagt accccaccat ctaccacctg agaaagaaac tggtggacag
1801 caccgacaag gccgacctgc ggctgatcta tctggccctg gcccacatga tcaagttccg
1861 gggccacttc ctgatcgagg gcgacctgaa ccccgacaac agcgacgtgg acaagctgtt
1921 catccagctg gtgcagacct acaaccagct gttcgaggaa aaccccatca acgccagcgg
1981 cgtggacgcc aaggccatcc tgtctgccag actgagcaag agcagacggc tggaaaatct
2041 gatcgcccag ctgcccggcg agaagaagaa tggcctgttc ggaaacctga ttgccctgag
2101 cctgggcctg acccccaact tcaagagcaa cttcgacctg gccgaggatg ccaaactgca
2161 gctgagcaag gacacctacg acgacgacct ggacaacctg ctggcccaga tcggcgacca
2221 gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac gccatcctgc tgagcgacat
2281 cctgagagtg aacaccgaga tcaccaaggc ccccctgagc gcctctatga tcaagagata
2341 cgacgagcac caccaggacc tgaccctgct gaaagctctc gtgcggcagc agctgcctga
2401 gaagtacaaa gagattttct tcgaccagag caagaacggc tacgccggct acattgacgg
2461 cggagccagc caggaagagt tctacaagtt catcaagccc atcctggaaa agatggacgg
2521 caccgaggaa ctgctcgtga agctgaacag agaggacctg ctgcggaagc agcggacctt
2581 cgacaacggc agcatccccc accagatcca cctgggagag ctgcacgcca ttctgcggcg
2641 gcaggaagat ttttacccat tcctgaagga caaccgggaa aagatcgaga agatcctgac
2701 cttccgcatc ccctactacg tgggccctct ggccagggga aacagcagat tcgcctggat
2761 gaccagaaag agcgaggaaa ccatcacccc ctggaacttc gaggaagtgg tggacaaggg
2821 cgcttccgcc cagagcttca tcgagcggat gaccaacttc gataagaacc tgcccaacga
2881 gaaggtgctg cccaagcaca gcctgctgta cgagtacttc accgtgtata acgagctgac
2941 caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc ttcctgagcg gcgagcagaa
3001 aaaggccatc gtggacctgc tgttcaagac caaccggaaa gtgaccgtga agcagctgaa
3061 agaggactac ttcaagaaaa tcgagtgctt cgactccgtg gaaatctccg gcgtggaaga
3121 tcggttcaac gcctccctgg gcacatacca cgatctgctg aaaattatca aggacaagga
3181 cttcctggac aatgaggaaa acgaggacat tctggaagat atcgtgctga ccctgacact
3241 gtttgaggac agagagatga tcgaggaacg gctgaaaacc tatgcccacc tgttcgacga
3301 caaagtgatg aagcagctga agcggcggag atacaccggc tggggcaggc tgagccggaa
3361 gctgatcaac ggcatccggg acaagcagtc cggcaagaca atcctggatt tcctgaagtc
3421 cgacggcttc gccaacagaa acttcatgca gctgatccac gacgacagcc tgacctttaa
3481 agaggacatc cagaaagccc aggtgtccgg ccagggcgat agcctgcacg agcacattgc
3541 caatctggcc ggcagccccg ccattaagaa gggcatcctg cagacagtga aggtggtgga
3601 cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac atcgtgatcg aaatggccag
3661 agagaaccag accacccaga agggacagaa gaacagccgc gagagaatga agcggatcga
3721 agagggcatc aaagagctgg gcagccagat cctgaaagaa caccccgtgg aaaacaccca
3781 gctgcagaac gagaagctgt acctgtacta cctgcagaat gggcgggata tgtacgtgga
3841 ccaggaactg gacatcaacc ggctgtccga ctacgatgtg gaccatatcg tgcctcagag
3901 ctttctggcc gacgactcca tcgacaacaa ggtgctgacc agaagcgaca agaaccgggg
3961 caagagcgac aacgtgccct ccgaagaggt cgtgaagaag atgaagaact actggcggca
4021 gctgctgaac gccaagctga ttacccagag aaagttcgac aatctgacca aggccgagag
4081 aggcggcctg agcgaactgg ataaggccgg cttcatcaag agacagctgg tggaaacccg
4141 gcagatcaca aagcacgtgg cacagatcct ggactcccgg atgaacacta agtacgacga
4201 gaatgacaag ctgatccggg aagtgaaagt gatcaccctg aagtccaagc tggtgtccga
4261 tttccggaag gatttccagt tttacaaagt gcgcgagatc aacaactacc accacgccca
4321 cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc aaaaagtacc ctaagctgga
4381 aagcgagttc gtgtacggcg actacaaggt gtacgacgtg cggaagatga tcgccaagag
4441 cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc tacagcaaca tcatgaactt
4501 tttcaagacc gagattaccc tggccaacgg cgagatccgg aagcggcctc tgatcgagac
4561 aaacggcgaa accggggaga tcgtgtggga taagggccgg gattttgcca ccgtgcggaa
4621 agtgctgagc atgccccaag tgaatatcgt gaaaaagacc gaggtgcaga caggcggctt
4681 cagcaaagag tctatcctgc ccaagaggaa cagcgataag ctgatcgcca gaaagaagga
4741 ctgggaccct aagaagtacg gcggcttcga cagccccacc gtggcctatt ctgtgctggt
4801 ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag agtgtgaaag agctgctggg
4861 gatcaccatc atggaaagaa gcagcttcga gaagaatccc atcgactttc tggaagccaa
4921 gggctacaaa gaagtgaaaa aggacctgat catcaagctg cctaagtact ccctgttcga
4981 gctggaaaac ggccggaaga gaatgctggc ctctgccggc gaactgcaga agggaaacga
5041 actggccctg ccctccaaat atgtgaactt cctgtacctg gccagccact atgagaagct
5101 gaagggctcc cccgaggata atgagcagaa acagctgttt gtggaacagc acaagcacta
5161 cctggacgag atcatcgagc agatcagcga gttctccaag agagtgatcc tggccgacgc
5221 taatctggac aaagtgctgt ccgcctacaa caagcaccgg gataagccca tcagagagca
5281 ggccgagaat atcatccacc tgtttaccct gaccaatctg ggagcccctg ccgccttcaa
5341 gtactttgac accaccatcg accggaagag gtacaccagc accaaagagg tgctggacgc
5401 caccctgatc caccagagca tcaccggcct gtacgagaca cggatcgacc tgtctcagct
5461 gggaggcgac aaaaggccgg cggccacgaa aaaggccggc caggcaaaaa agaaaaagga
5521 attcggcagt ggagagggca gaggaagtct gctaacatgc ggtgacgtcg aggagaatcc
5581 tggcccaatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
5641 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
5701 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
5761 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
5821 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
5881 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
5941 caaggagccc gcgtggttcc tggccaccgt cggagtctcg cccgaccacc agggcaaggg
6001 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
6061 cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
6121 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
6181 tgcctgagaa ttctaactag agctcgctga tcagcctcga ctgtgccttc tagttgccag
6241 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
6301 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
6361 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagagaa tagcaggcat
6421 gctggggagc ggccgcagga acccctagtg atggagttgg ccactccctc tctgcgcgct
6481 cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg
6541 gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc
6601 cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct
6661 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
6721 ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
6781 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
6841 ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct
6901 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
6961 tccaaactgg aacaacactc aactctatct cgggctattc ttttgattta taagggattt
7021 tgccgatttc ggtctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
7081 ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg
7141 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
7201 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
7261 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat
7321 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
7381 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
7441 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
7501 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
7561 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
7621 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
7681 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
7741 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
7801 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
7861 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
7921 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
7981 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
8041 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
8101 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
8161 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtgga agccgcggta
8221 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
8281 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
8341 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
8401 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
8461 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
8521 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
8581 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
8641 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
8701 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
8761 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
8821 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
8881 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
8941 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
9001 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
9061 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
9121 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgt
//
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