| 质粒名称: | pNICKclos2.0 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 73228 |
| 存储实验室: | Sheng Yang |
|
载体骨架基本信息 |
|
| 载体名称: | pXY1 |
| 载体出品公司: | -- |
| 载体抗性: | low Amp |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | E、 大肠杆菌梭状芽孢杆菌穿梭载体 |
| 筛选标记: | 梭状芽孢杆菌红霉素 |
| 高拷贝/低拷贝: | 低拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | Cas9 nickase |
| 物种来源: | S. pyogenes MGAS5005 |
| 插入大小: | -- |
| 基因突变: | D10A |
| GenBank编号 | -- |
| 启动子 | Pthl |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | PstI(未破坏) |
| 3' 克隆位点: | NotI (未破坏) |
| 5' 测序引物 | SpCas9N-R TACGAGCTGTCCGTTTGAG |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pNICKclos2.0 was a gift from Sheng Yang (Addgene plasmid # 73228 ; http://n2t.net/addgene:73228 ; RRID:Addgene_73228)A |
| 参考文献部分: | CRISPR-based genome editing and expression control systems in Clostridium acetobutylicum and Clostridium beijerinckii. Li Q, Chen J, Minton NP, Zhang Y, Wen Z, Liu J, Yang H, Zeng Z, Ren X, Yang J, Gu Y, Jiang W, Jiang Y, Yang S. Biotechnol J. 2016 May 23. doi: 10.1002/biot.201600053. 10.1002/biot.201600053 PubMed 27213844 |
买家导航
pNICKclos2.0载体质粒基本信息
pNICKclos2.0质粒图谱载体图谱和pNICKclos2.0载体序列质粒序列多克隆位点信息
pNICKclos2.0质粒载体简介
pNICKclos2.0质粒序列载体序列
LOCUS pNICKclos2.0 11706 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Genome editing for gene xylR (cbei-2385) in clostridium beijerinckii
NCIMB 8052.
ACCESSION .
VERSION .
KEYWORDS pNICKclos2.0
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 11706)
AUTHORS Li Q, Chen J, Minton NP, Zhang Y, Wen Z, Liu J, Yang H, Zeng Z, Ren
X, Yang J, Gu Y, Jiang W, Jiang Y, Yang S
TITLE CRISPR-based genome editing and expression control systems in
Clostridium acetobutylicum and Clostridium beijerinckii.
JOURNAL Biotechnol J. 2016 May 23. doi: 10.1002/biot.201600053.
PUBMED 27213844
REFERENCE 2 (bases 1 to 11706)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..11706
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1488..1522
/label=J23119(SpeI) promoter
/note="bacterial promoter (Registry of Standard Biological
Parts BBa_J23119) modified to end with an SpeI site"
misc_RNA 1543..1618
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
CDS 3804..7910
/codon_start=1
/product="nickase mutant of the Cas9 endonuclease from the
Streptococcus pyogenes Type II CRISPR/Cas system"
/label=Cas9(D10A)
/note="generates RNA-guided single strand nicks in DNA due
to the D10A mutation in the RuvC catalytic domain"
/translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
primer_bind complement(7927..7944)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(7927..7943)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(7936..7958)
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind complement(8152..8171)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 8271..8293
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(8331..8349)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 8417..8521
/gene="bla"
/label=AmpR promoter
CDS 8522..9382
/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(8740..8759)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 9553..10141
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 10042..10061
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 10295..10312
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 10429..10450
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 10465..10495
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 10503..10519
/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 10508..10530
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 10527..10543
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 10527..10543
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
ORIGIN
1 cgataattta cagaaaagaa aattatagaa tttagtatga ttaattatac tcatttatga
61 atgtttaatt gaatacaaaa aaaaantact tgttatgtat tcaattacgg gttaaaatat
121 agacaagttg aaaaatttaa taaaaaaata agtcctcagc tcttatatat taagctacca
181 acttagtata taagccaaaa cttaaatgtg ctaccaacac atcaagccgt tagagaactc
241 tatctatagc aatatttcaa atgtaccgac atacaagaga aacattaact atatatattc
301 aatttatgag attatcttaa cagatataaa tgtaaattgc aataagtaag atttagaagt
361 ttatagcctt tgtgtattgg aagcagtacg caaaggcttt tttatttgat aaaaattaga
421 agtatattta ttttttcata attaatttat gaaaatgaaa gggggtgagc aaagtgacag
481 aggaaagcag tatcttatca aataacaagg tattagcaat atcattattg actttagcag
541 taaacattat gacttttata gtgcttgtag ctaagtagta cgaaaggggg agctttaaaa
601 agctccttgg aatacataga attcataaat taatttatga aaagaagggc gtatatgaaa
661 acttgtaaaa attgcaaaga gtttattaaa gatactgaaa tatgcaaaat acattcgttg
721 atgattcatg ataaaacagt agcaacctat tgcagtaaat acaatgagtc aagatgttta
781 cataaaggga aagtccaatg tattaattgt tcaaagatga accgatatgg atggtgtgcc
841 ataaaaatga gatgttttac agaggaagaa cagaaaaaag aacgtacatg cattaaatat
901 tatgcaagga gctttaaaaa agctcatgta aagaagagta aaaagaaaaa ataatttatt
961 tattaattta atattgagag tgccgacaca gtatgcacta aaaaatatat ctgtggtgta
1021 gtgagccgat acaaaaggat agtcactcgc attttcataa tacatcttat gttatgatta
1081 tgtgtcggtg ggacttcacg acgaaaaccc acaataaaaa aagagttcgg ggtagggtta
1141 agcatagttg aggcaactaa acaatcaagc taggatatgc agtagcagac cgtaaggtcg
1201 ttgtttaggt gtgttgtaat acatacgcta ttaagatgta aaaatacgga taccaatgaa
1261 gggaaaagta taatttttgg atgtagtttg tttgttcatc tatgggcaaa ctacgtccaa
1321 agccgtttcc aaatctgcta aaaagtatat cctttctaaa atcaaagtca agtatgaaat
1381 cataaataaa gtttaatttt gaagttatta tgatattatg tttttctatt aaaataaatt
1441 aagtatatag aatagtttaa taatagtata tacttaatgt gctgcagttg acagctagct
1501 cagtcctagg tataatacta gtcgagttag acataatagt gagttttaga gctagaaata
1561 gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt
1621 ttttgcggcc gcaaaacctg gttctaatgg aattgtttac ttaccatatc ttatgggaga
1681 aagaacacct catatagacc ctaatgtaaa aggtgcattt ttaggagtat cactaataaa
1741 taaccatgat gattttgtac gtagtatatt agaaggtgtt agctttagtt tgaaaaactg
1801 ccttgattta atcgaagaca tgaatgtaaa cattgatgaa ataagagtaa gtggcggtgg
1861 agcagaaagt gctgtatgga gacaaatatt agctgatatt ttccaatatt cattaactac
1921 agttaaagca tctgaaggcg gtgcgcttgg agttgcaata ttagcagggg ttggagcagg
1981 aatctatgat tctgtagaag atgcttgtga tagaatcgta aaaggaaaca agaaagtttc
2041 tccaaatgaa gaattaaaag aagtatactc aaaagtatat gaaacatata attcagctta
2101 tccaagaata aaagatattt aaatatatta aagatagaaa tcaatcatct tagttaagta
2161 attgtataat taattaaggg tgtgtagtag tatcgaattt tagcaaaaaa atcatatagt
2221 tttttaaata ctttaacaaa aataataata gtgatattat ttaagaaaaa gctactatta
2281 ctgataatag tcgagaggag tattttaaag tttgatagaa gtaaatcaca gtaaaataaa
2341 agaaactaac agaaaaaaga taatagcatt attattaaag aaaaatgaga tcaccaaatt
2401 agatatatct agaatattag atataagtat aacaacagtt tctacaaata ttacagaatt
2461 gaaaagcgaa ggtatnnnnn nnnatgtaag atcgttggaa tctacaggag gaagaaaagc
2521 aattgctatt aagttgaatg aaaactgtag atattcaatt ggagtcgcac taacaccgaa
2581 tcatataaag atatcattag taaatataaa gaaaaaagtt attgaaagta tgctgcagtg
2641 gaatagagaa cataataaat ttacttaatg aaaatataga tttgttaatg aagaaatata
2701 atttatgctc tgcaaattta ttaggaatag ggatttcatt acctggaacc gtagatttta
2761 aagaaggcat aataaagtat tcatatttgt taggcgccaa agattttaat ttaaaggaaa
2821 agttcgaata cttagacatt ccagtatatg tagataatga agcaaattta tctgcttact
2881 acgagttttt aaataaaaga gatatactaa gaaatctatt atatgtatca ataacagaag
2941 gtttaggcct tggtataata ataaatggaa aaatttacag aggtgacaat aactcatcag
3001 gagaattagg tcatactaag atagcaattg atggaaaaaa atgtaagtgt ggagctagag
3061 gctgtttaga agcatacaca tctatgaatt ctctgattga tagttataat gaagctaata
3121 gttcaaatat tagtgatatt gatgaatttg aagaaagata taatcaaaat gacaaagatg
3181 ctcatgatgt attaaatgat tatttaaaaa cattgtcttt aggtctatcg aatttggtta
3241 tgatatttga tcctagcagc atagtaatag gtggagatat aaataattta cttaatgata
3301 agatagatat gctgaaaaaa gaggtttata aagataattt atttacaaat gaaaataatt
3361 gcagtataag tattgcaagt tttaaagaat cttatctact aggagctgct atgatgccta
3421 tagaagaatt cttagaaatt aaataatgga gtggtgaaag ctgctctatt aatttaaaaa
3481 cttattaaat aaatttttaa aatatatttt taaagttatg taagcatagc tatattaagg
3541 ggggtcttaa tctttgattg gaactccaga ttaagacaag ccttttatat aaaactaaat
3601 atctatttat ttcatattag atttatgttc ttgattttct cgagttttta acaaaatata
3661 ttgataaaaa taataatagt gggtataatt aagttgttag agaaaacgta taaattaggg
3721 ataaactatg gaacttatga aatagattga aatggtttat ctgttacccc gtatcaaaat
3781 ttaggaggtt agttagaccc gggatggata agaaatactc aataggctta gctatcggca
3841 caaatagcgt cggatgggcg gtgatcactg atgaatataa ggttccgtct aaaaagttca
3901 aggttctggg aaatacagac cgccacagta tcaaaaaaaa tcttataggg gctcttttat
3961 ttgacagtgg agagacagcg gaagcgactc gtctcaaacg gacagctcgt agaaggtata
4021 cacgtcggaa gaatcgtatt tgttatctac aggagatttt ttcaaatgag atggcgaaag
4081 tagatgatag tttctttcat cgacttgaag agtctttttt ggtggaagaa gacaagaagc
4141 atgaacgtca tcctattttt ggaaatatag tagatgaagt tgcttatcat gagaaatatc
4201 caactatcta tcatctgcga aaaaaattgg tagattctac tgataaagcg gatttgcgct
4261 taatctattt ggccttagcg catatgatta agtttcgtgg tcattttttg attgagggag
4321 atttaaatcc tgataatagt gatgtggaca aactatttat ccagttggta caaacctaca
4381 atcaattatt tgaagaaaac cctattaacg caagtggagt agatgctaaa gcgattcttt
4441 ctgcacgatt gagtaaatca agacgattag aaaatctcat tgctcagctc cccggtgaga
4501 agaaaaatgg cttatttggg aatctcattg ctttgtcatt gggtttgacc cctaatttta
4561 aatcaaattt tgatttggca gaagatgcta aattacagct ttcaaaagat acttacgatg
4621 atgatttaga taatttattg gcgcaaattg gagatcaata tgctgatttg tttttggcag
4681 ctaagaattt atcagatgct attttacttt cagatatcct aagagtaaat actgaaataa
4741 ctaaggctcc cctatcagct tcaatgatta aacgctacga tgaacatcat caagacttga
4801 ctcttttaaa agctttagtt cgacaacaac ttccagaaaa gtataaagaa atcttttttg
4861 atcaatcaaa aaacggatat gcaggttata ttgatggggg agctagccaa gaagaatttt
4921 ataaatttat caaaccaatt ttagaaaaaa tggatggtac tgaggaatta ttggtgaaac
4981 taaatcgtga agatttgctg cgcaagcaac ggacctttga caacggctct attccccatc
5041 aaattcactt gggtgagctg catgctattt tgagaagaca agaagacttt tatccatttt
5101 taaaagacaa tcgtgagaag attgaaaaaa tcttgacttt tcgaattcct tattatgttg
5161 gtccattggc gcgtggcaat agtcgttttg catggatgac tcggaagtct gaagaaacaa
5221 ttaccccatg gaattttgaa gaagttgtcg ataaaggtgc ttcagctcaa tcatttattg
5281 aacgcatgac aaactttgat aaaaatcttc caaatgaaaa agtactacca aaacatagtt
5341 tgctttatga gtattttacg gtttataacg aattgacaaa ggtcaaatat gttactgaag
5401 gaatgcgaaa accagcattt ctttcaggtg aacagaagaa agccattgtt gatttactct
5461 tcaaaacaaa tcgaaaagta accgttaagc aattaaaaga agattatttc aaaaaaatag
5521 aatgttttga tagtgttgaa atttcaggag ttgaagatag atttaatgct tcattaggta
5581 cctaccatga tttgctaaaa attattaaag ataaagattt tttggataat gaagaaaatg
5641 aagatatctt agaggatatt gttttaacat tgaccttatt tgaagatagg gagatgattg
5701 aggaaagact taaaacatat gctcacctct ttgatgataa ggtgatgaaa cagcttaaac
5761 gtcgccgtta tactggttgg ggacgtttgt ctcgaaaatt gattaatggt attagggata
5821 agcaatctgg caaaacaata ttagattttt tgaaatcaga tggttttgcc aatcgcaatt
5881 ttatgcagct gatccatgat gatagtttga catttaaaga agacattcaa aaagcacaag
5941 tgtctggaca aggcgatagt ttacatgaac atattgcaaa tttagctggt agccctgcta
6001 ttaaaaaagg tattttacag actgtaaaag ttgttgatga attggtcaaa gtaatggggc
6061 ggcataagcc agaaaatatc gttattgaaa tggcacgtga aaatcagaca actcaaaagg
6121 gccagaaaaa ttcgcgagag cgtatgaaac gaatcgaaga aggtatcaaa gaattaggaa
6181 gtcagattct taaagagcat cctgttgaaa atactcaatt gcaaaatgaa aagctctatc
6241 tctattatct ccaaaatgga agagacatgt atgtggacca agaattagat attaatcgtt
6301 taagtgatta tgatgtcgat cacattgttc cacaaagttt ccttaaagac gattcaatag
6361 acaataaggt cttaacgcgt tctgataaaa atcgtggtaa atcggataac gttccaagtg
6421 aagaagtagt caaaaagatg aaaaactatt ggagacaact tctaaacgcc aagttaatca
6481 ctcaacgtaa gtttgataat ttaacgaaag ctgaacgtgg aggtttgagt gaacttgata
6541 aagctggttt tatcaaacgc caattggttg aaactcgcca aatcactaag catgtggcac
6601 aaattttgga tagtcgcatg aatactaaat acgatgaaaa tgataaactt attcgagagg
6661 ttaaagtgat taccttaaaa tctaaattag tttctgactt ccgaaaagat ttccaattct
6721 ataaagtacg tgagattaac aattaccatc atgcccatga tgcgtatcta aatgccgtcg
6781 ttggaactgc tttgattaag aaatatccaa aacttgaatc ggagtttgtc tatggtgatt
6841 ataaagttta tgatgttcgt aaaatgattg ctaagtctga gcaagaaata ggcaaagcaa
6901 ccgcaaaata tttcttttac tctaatatca tgaacttctt caaaacagaa attacacttg
6961 caaatggaga gattcgcaaa cgccctctaa tcgaaactaa tggggaaact ggagaaattg
7021 tctgggataa agggcgagat tttgccacag tgcgcaaagt attgtccatg ccccaagtca
7081 atattgtcaa gaaaacagaa gtacagacag gcggattctc caaggagtca attttaccaa
7141 aaagaaattc ggacaagctt attgctcgta aaaaagactg ggatccaaaa aaatatggtg
7201 gttttgatag tccaacggta gcttattcag tcctagtggt tgctaaggtg gaaaaaggga
7261 aatcgaagaa gttaaaatcc gttaaagagt tactagggat cacaattatg gaaagaagtt
7321 cctttgaaaa aaatccgatt gactttttag aagctaaagg atataaggaa gttaaaaaag
7381 acttaatcat taaactacct aaatatagtc tttttgagtt agaaaacggt cgtaaacgga
7441 tgctggctag tgccggagaa ttacaaaaag gaaatgagct ggctctgcca agcaaatatg
7501 tgaatttttt atatttagct agtcattatg aaaagttgaa gggtagtcca gaagataacg
7561 aacaaaaaca attgtttgtg gagcagcata agcattattt agatgagatt attgagcaaa
7621 tcagtgaatt ttctaagcgt gttattttag cagatgccaa tttagataaa gttcttagtg
7681 catataacaa acatagagac aaaccaatac gtgaacaagc agaaaatatt attcatttat
7741 ttacgttgac gaatcttgga gctcccgctg cttttaaata ttttgataca acaattgatc
7801 gtaaacgata tacgtctaca aaagaagttt tagatgccac tcttatccat caatccatca
7861 ctggtcttta tgaaacacgc attgatttga gtcagctagg aggtgactga gcggccgcgg
7921 gaattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
7981 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
8041 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga tgcggtattt
8101 tctccttacg catctgtgcg gtatttcaca ccgcatatgg tgcactctca gtacaatctg
8161 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
8221 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
8281 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
8341 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
8401 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
8461 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
8521 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
8581 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
8641 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
8701 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
8761 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
8821 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
8881 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
8941 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
9001 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
9061 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
9121 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
9181 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
9241 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
9301 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
9361 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
9421 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
9481 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
9541 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
9601 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
9661 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
9721 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
9781 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
9841 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
9901 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
9961 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
10021 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
10081 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
10141 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
10201 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
10261 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
10321 agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
10381 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
10441 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
10501 aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagct
10561 ttggctaaca cacacgccat tccaaccaat agttttctcg gcataaagcc atgctctgac
10621 gcttaaatgc actaatgcct taaaaaaaca ttaaagtcta acacactaga cttatttact
10681 tcgtaattaa gtcgttaaac cgtgtgctct acgaccaaaa gtataaaacc tttaagaact
10741 ttcttttttc ttgtaaaaaa agaaactaga taaatctctc atatctttta ttcaataatc
10801 gcatcagatt gcagtataaa tttaacgatc actcatcatg ttcatattta tcagagctcg
10861 tgctataatt atactaattt tataaggagg aaaaaataaa gagggttata atgaacgaga
10921 aaaatataaa acacagtcaa aactttatta cttcaaaaca taatatagat aaaataatga
10981 caaatataag attaaatgaa catgataata tctttgaaat cggctcagga aaagggcatt
11041 ttacccttga attagtacag aggtgtaatt tcgtaactgc cattgaaata gaccataaat
11101 tatgcaaaac tacagaaaat aaacttgttg atcacgataa tttccaagtt ttaaacaagg
11161 atatattgca gtttaaattt cctaaaaacc aatcctataa aatatttggt aatatacctt
11221 ataacataag tacggatata atacgcaaaa ttgtttttga tagtatagct gatgagattt
11281 atttaatcgt ggaatacggg tttgctaaaa gattattaaa tacaaaacgc tcattggcat
11341 tatttttaat ggcagaagtt gatatttcta tattaagtat ggttccaaga gaatattttc
11401 atcctaaacc taaagtgaat agctcactta tcagattaaa tagaaaaaaa tcaagaatat
11461 cacacaaaga taaacagaag tataattatt tcgttatgaa atgggttaac aaagaataca
11521 agaaaatatt tacaaaaaat caatttaaca attccttaaa acatgcagga attgacgatt
11581 taaacaatat tagctttgaa caattcttat ctcttttcaa tagctataaa ttatttaata
11641 agtaagttaa gggatgcata aactgcatcc cttaacttgt ttttcgtgta cctatttttt
11701 gtgaat
//
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