| 质粒名称: | pJH2971 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 100955 |
| 存储实验室: | James Haber |
|
载体骨架基本信息 |
|
| 载体名称: | pRS316 |
| 载体出品公司: | NA |
| 完整质粒大小: | 11162bp |
| 载体类型: | Yeast Expression, CRISPR, Synthetic Biology |
| 筛选标记: | 新霉素（G418） |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | 无 |
|
文献引用方法 |
|
| 材料和方法部分: | pJH2971 was a gift from James Haber (Addgene plasmid # 100955 ; http://n2t.net/addgene:100955 ; RRID:Addgene_100955) |
| 参考文献部分: | Cas9-mediated gene editing in Saccharomyces cerevisiae. Ranjith Anand, Gonen Memisoglu & James Haber. Protocol Exchange (2017) 10.1038/protex.2017.021a |
买家导航
pJH2971载体质粒基本信息
pJH2971质粒图谱载体图谱和pJH2971载体序列质粒序列多克隆位点信息
pJH2971质粒载体简介
pJH2971质粒序列载体序列
LOCUS pJH2971 11162 bp DNA circular SYN 14-APR-2025
DEFINITION Cas9 driven by PGK1 promoter. gRNA can be cloned into the BplI
sites. KANMX marker.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 11162)
AUTHORS
TITLE Cas9-mediated gene editing in Saccharomyces cerevisiae
JOURNAL Protocol Exchange (2017)
REFERENCE 2 (bases 1 to 11162)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 14, 2025 from SnapGene Server 8.0.1
https://www.biofeng.com
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Protocol
Exchange (2017)"
COMMENT Sequence Label: pJH2971
FEATURES Location/Qualifiers
source 1..11162
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind complement(29..51)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 151..170
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
gene complement(284..1640)
/label=kanMX
/note="yeast selectable marker conferring kanamycin
resistance (Wach et al., 1994)"
terminator complement(284..481)
/label=TEF terminator
/note="Ashbya gossypii TEF terminator"
CDS complement(487..1296)
/codon_start=1
/gene="aph(3')-Ia"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin"
/translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
primer_bind 1210..1229
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
promoter complement(1297..1640)
/label=TEF promoter
/note="Ashbya gossypii TEF promoter"
CDS 2658..6761
/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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
CDS 2658..6761
/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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
CDS 6774..6794
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 6798..6818
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 6822..6842
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 6843..6908
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
terminator 6927..7174
/gene="S. cerevisiae CYC1"
/label=CYC1 terminator
/note="transcription terminator for CYC1"
primer_bind complement(6943..6961)
/label=CYC1
/note="CYC1 transcription termination signal, reverse
primer"
rep_origin complement(7234..7689)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(7371..7392)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 7583..7602
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 7815..7837
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 7829..7846
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 7830..7846
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 7853..7872
/label=T7
/note="T7 promoter, forward primer"
promoter 7853..7871
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind 7904..7920
/label=SK primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 7904..7920
/label=pBluescriptSK
/note="For pBluescript vector"
promoter 7952..8220
/label=SNR52 promoter
/note="promoter for the S. cerevisiae small nucleolar RNA
gene SNR52"
misc_RNA 8248..8322
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
terminator 8327..8346
/label=SUP4 terminator
/note="transcription terminator for the S. cerevisiae SUP4
tRNA gene"
promoter complement(8389..8407)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(8428..8444)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(8428..8444)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(8441..8463)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 8452..8468
/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(join(8476..8482,8483..8500,8501..8506))
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 8521..8542
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(8659..8676)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(8830..9418)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(8910..8929)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(join(9589..10380,10381..10449))
/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 10212..10231
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(10450..10554)
/gene="bla"
/label=AmpR promoter
misc_feature 10591..11094
/label=CEN/ARS
/note="S. cerevisiae CEN6 centromere fused to an
autonomously replicating sequence"
primer_bind 11135..11153
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
ORIGIN
1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
181 accataccac cttttcaatt catcattttt tttttattct tttttttgat ttcgaattcg
241 agctcgttta aactggatgg cggcgttagt atcgaatcga cagcagtata gcgaccagca
301 ttcacatacg attgacgcat gatattactt tctgcgcact taacttcgca tctgggcaga
361 tgatgtcgag gcgaaaaaaa atataaatca cgctaacatt tgattaaaat agaacaacta
421 caatataaaa aaactataca aatgacaagt tcttgaaaac aagaatcttt ttattgtcag
481 tactgattag aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt
541 atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca
601 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
661 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
721 gacgactgaa tccggtgaga atggcaaaag cttatgcatt tctttccaga cttgttcaac
781 aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg
841 tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg
901 aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
961 aggatattct tctaatacct ggaatgctgt tttgccgggg atcgcagtgg tgagtaacca
1021 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag
1081 ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
1141 cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg
1201 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
1261 tcgcggcctc gaaacgtgag tcttttcctt acccatggtt gtttatgttc ggatgtgatg
1321 tgagaactgt atcctagcaa gattttaaaa ggaagtatat gaaagaagaa cctcagtggc
1381 aaatcctaac cttttatatt tctctacagg ggcgcggcgt ggggacaatt caacgcgtct
1441 gtgaggggag cgtttccctg ctcgcaggtc cgcagcgagg agccgtaatt tttgcttcgc
1501 gccgtgcggc catcaaaatg tatggatgca aatgattata catggggatg tatgggctaa
1561 atgtacgggc gacagtcaca tcatgcccct gagctgcgca cgtcaagact gtcaaggagg
1621 gtattctggg cctccatgtc gctggccggg tgacccggcg gggaaagtac cttcaaagaa
1681 tggggtctta tcttgttttg caagtaccac tgagcaggat aataatagaa atgataatat
1741 actatagtag agataacgtc gatgacttcc catactgtaa ttgcttttag ttgtgtattt
1801 ttagtgtgca agtttctgta aatcgattaa tttttttttc tttcctcttt ttattaacct
1861 taatttttat tttagattcc tgacttcaac tcaagacgca cagatattat aacatctgca
1921 taataggcat ttgcaagaat tactcgtgag taaggaaaga gtgaggaact atcgcatacc
1981 tgcatttaaa gatgccgatt tgggcgcgaa tcctttattt tggcttcacc ctcatactat
2041 tatcagggcc agaaaaagga agtgtttccc tccttcttga attgatgtta ccctcataaa
2101 gcacgtggcc tcttatcgag aaagaaatta ccgtcgctcg tgatttgttt gcaaaaagaa
2161 caaaactgaa aaaacccaga cacgctcgac ttcctgtctt cctattgatt gcagcttcca
2221 atttcgtcac acaacaaggt cctagcgacg gctcacaggt tttgtaacaa gcaatcgaag
2281 gttctggaat ggcgggaaag ggtttagtac cacatgctat gatgcccact gtgatctcca
2341 gagcaaagtt cgttcgatcg tactgttact ctctctcttt caaacagaat tgtccgaatc
2401 gtgtgacaac aacagcctgt tctcacacac tcttttcttc taaccaaggg ggtggtttag
2461 tttagtagaa cctcgtgaaa cttacattta catatatata aacttgcata aattggtcaa
2521 tgcaagaaat acatatttgg tcttttctaa ttcgtagttt ttcaagttct tagatgcttt
2581 ctttttctct tttttacaga tcatcaagga agtaattatc tactttttac aacaaatata
2641 aaacaccgga tctcaaaatg gataaaaagt atagtattgg tttagatatt ggtactaact
2701 ctgtgggttg ggcagttatc actgacgaat ataaagttcc atcaaagaaa tttaaggtgt
2761 taggtaacac tgacagacac tcaataaaaa agaatcttat cggtgctctt ttgttcgact
2821 ccggtgaaac tgccgaggct acacgtttaa aaagaacagc aagaagaaga tatacccgta
2881 gaaaaaatag aatatgttat ttacaagaaa tcttttctaa tgaaatggct aaagttgatg
2941 attccttttt ccatagattg gaagagtcat ttttggttga agaagacaaa aagcatgaga
3001 gacatccaat ctttgggaat atagttgatg aagtggctta ccatgaaaaa tatcctacca
3061 tttatcattt aagaaagaaa ttggtagatt caactgataa agctgacctt agattaatct
3121 atttagcact tgcccatatg attaaattta gaggtcattt tttgattgaa ggtgatttga
3181 acccagataa ttctgacgtg gataaattat ttattcaatt agtccaaacc tacaaccaat
3241 tatttgagga aaatccaatt aatgctagtg gtgtcgatgc caaagctata ttatcagcca
3301 gattatcaaa atctagacgt ttggaaaatt tgattgccca attgccagga gaaaaaaaga
3361 atggattatt tggaaacttg atcgcattat cattgggttt gacaccaaat tttaaatcta
3421 attttgattt agctgaagat gctaaattac aattatcaaa agacacctat gacgacgatt
3481 tggacaattt acttgctcaa attggtgatc aatatgcaga tttgttctta gctgctaaaa
3541 acttatctga tgctattttg ttgtctgata ttttgagagt gaacacagaa ataaccaaag
3601 ctccattatc agcatctatg atcaaacgtt atgatgaaca ccatcaggat ttgactttat
3661 tgaaagcttt ggtgagacaa caattgccag agaagtataa agaaatcttt ttcgatcaat
3721 ctaaaaacgg gtatgcaggt tatattgatg ggggtgcctc ccaagaggaa ttttacaaat
3781 ttataaaacc tattttagaa aagatggatg ggactgagga acttttggtc aaattgaaca
3841 gagaagattt gttacgtaaa cagagaactt ttgataatgg tagtatacct caccaaattc
3901 atttgggtga gttgcatgca attttaagaa gacaagaaga tttttatcca tttttaaaag
3961 ataatagaga aaaaatcgag aaaattttaa cctttagaat tccatactat gttgggcctt
4021 tggctagagg taattcaaga tttgcctgga tgacacgtaa atcagaagaa actattaccc
4081 cttggaattt tgaagaggtt gttgataaag gagcatcagc acagagtttt attgaaagaa
4141 tgaccaattt cgataaaaac ttaccaaatg aaaaagtttt accaaaacat tccttgttat
4201 acgaatattt tactgtttac aatgaactta caaaggttaa atatgttact gaaggtatgc
4261 gtaagccagc ctttttatct ggagaacaga aaaaggcaat agttgattta ttgtttaaaa
4321 caaatagaaa agttactgtt aaacaattaa aagaagatta ctttaagaaa attgaatgtt
4381 ttgattcagt tgaaatcagt ggtgttgaag acagatttaa tgctagttta ggaacttacc
4441 atgatttact taaaattatc aaagataaag atttcttgga taacgaagaa aatgaagaca
4501 ttttagaaga cattgtttta accttaactt tattcgaaga tagagagatg attgaagaac
4561 gtttgaagac ttatgcacat ttgtttgacg ataaagtgat gaaacagttg aaaagaagac
4621 gttatactgg atggggtaga ttgtctcgta aattgatcaa tggaattaga gataaacaaa
4681 gtggtaaaac tatcttggac tttttgaaat ctgacggatt tgctaataga aatttcatgc
4741 aattgatcca cgacgatagt ttgacattta aagaagacat ccaaaaggcc caagtgagtg
4801 ggcaaggtga ttcattacat gaacatattg caaatttagc cggatctcct gctattaaga
4861 aagggatatt acaaactgtt aaagttgtgg atgaattagt gaaagtaatg ggaagacata
4921 aacctgaaaa cattgtcatt gagatggcaa gagaaaatca aactacacaa aaaggacaga
4981 aaaatagtag agaacgtatg aaaagaatag aagagggtat taaagaattg ggtagtcaaa
5041 tattgaaaga acacccagtg gaaaataccc agttgcaaaa tgaaaaatta tatctttact
5101 accttcaaaa tggacgtgat atgtatgttg atcaggaatt agatataaat agactttcag
5161 attatgatgt agatcatata gttccacaat ctttcttgaa agatgattcc atagacaata
5221 aagtattaac tagaagtgat aaaaatagag gtaaaagtga taatgtccca agtgaggaag
5281 tcgtcaaaaa gatgaaaaat tactggcgtc aacttttgaa tgctaaatta attactcaaa
5341 gaaaatttga taatttgact aaagcagaaa gaggtgggct ttctgaatta gataaagccg
5401 ggttcattaa aagacaattg gtcgaaacta gacaaattac taaacatgtt gcccaaattt
5461 tagattcccg tatgaacact aagtatgacg aaaatgataa gttaatacgt gaggttaaag
5521 tcattacttt aaaatcaaaa cttgtctctg atttcagaaa ggatttccaa ttctataaag
5581 ttagagaaat taataattat catcatgctc atgatgcata tttgaatgct gtagttggaa
5641 ctgctttaat caagaaatac cctaaattag aatctgaatt tgtatatggt gattacaaag
5701 tctatgatgt tagaaagatg attgctaaat cagaacaaga aattggtaaa gctacagcta
5761 aatacttctt ttactctaac attatgaatt tctttaaaac agaaattact ttggcaaacg
5821 gtgaaattag aaaaagacct cttattgaaa caaatggtga gactggagag atagtttggg
5881 acaaagggcg tgatttcgct actgttagaa aagttttatc aatgccacaa gttaacattg
5941 taaagaaaac agaggttcaa actggtggtt tctcaaaaga aagtattttg cctaaaagaa
6001 atagtgataa attgattgcc agaaaaaagg attgggatcc aaagaaatat ggtggtttcg
6061 actcaccaac cgtagcctat tctgttttgg ttgtggcaaa ggttgaaaag ggtaaaagta
6121 aaaagcttaa atcagtaaaa gaacttttgg gtattacaat aatggaaaga agttcctttg
6181 aaaagaaccc tattgatttt ttggaagcta aaggttataa ggaagtaaag aaggacttaa
6241 taatcaaatt gcctaaatat tctttatttg aattagaaaa tgggagaaaa agaatgttgg
6301 cttctgctgg agaattgcaa aagggtaatg aattagcatt gccttccaaa tatgttaact
6361 tcttgtattt agcttcacac tatgaaaagt tgaaagggtc accagaagat aacgagcaaa
6421 aacaattatt tgttgaacaa cacaaacact acttagatga gattatagaa caaattagtg
6481 aattcagtaa aagagtgata ttagctgatg caaatttaga taaagttttg tcagcctata
6541 acaaacatag agataagcca attagagaac aagcagaaaa cattattcac ttatttaccc
6601 ttaccaattt aggagcacct gctgctttca agtattttga tacaacaatt gatcgtaaaa
6661 gatatacctc aacaaaagaa gtcttagacg ccaccttaat tcatcaatca atcactggat
6721 tgtatgagac aagaattgat ttgtctcaat tgggtggtga tgaaggggct gatcctaaga
6781 agaaaagaaa agttgatcca aagaaaaagc gtaaggtgga tcctaagaaa aagagaaagg
6841 ttgactacaa agaccatgac ggtgattata aagatcatga catcgactac aaggatgacg
6901 atgacaagtg ataatctctt ctcgagtcat gtaattagtt atgtcacgct tacattcacg
6961 ccctcccccc acatccgctc taaccgaaaa ggaaggagtt agacaacctg aagtctaggt
7021 ccctatttat ttttttatag ttatgttagt attaagaacg ttatttatat ttcaaatttt
7081 tctttttttt ctgtacagac gcgtgtacgc atgtaacatt atactgaaaa ccttgcttga
7141 gaaggttttg ggacgctcga aggctttaat ttgcggccgg tactgcggtg tgaaataccg
7201 cacagatgcg taaggagaaa ataccgcatc aggaaattgt aagcgttaat attttgttaa
7261 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
7321 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
7381 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
7441 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
7501 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
7561 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
7621 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
7681 agggcgcgtc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
7741 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt
7801 aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgaat tgtaatacga
7861 ctcactatag ggcgaattgg agctccaccg cggtggcggc cgctctagaa ctagtggatc
7921 ccccgggctg caggaattcg atatcaagct ttctttgaaa agataatgta tgattatgct
7981 ttcactcata tttatacaga aacttgatgt tttctttcga gtatatacaa ggtgattaca
8041 tgtacgtttg aagtacaact ctagattttg tagtgccctc ttgggctagc ggtaaaggtg
8101 cgcatttttt cacaccctac aatgttctgt tcaaaagatt ttggtcaaac gctgtagaag
8161 tgaaagttgg tgcgcatgtt tcggcgttcg aaacttctcc gcagtgaaag ataaatgatc
8221 ctagatgcga gatatcctca tgcctaggtt ttagagctag aaatagcaag ttaaaataag
8281 gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tggtgctttt tttgtttttt
8341 atgtctaccg tcgacctcga gggggggccc ggtacccagc ttttgttccc tttagtgagg
8401 gttaatttcg agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
8461 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
8521 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
8581 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
8641 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
8701 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
8761 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
8821 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
8881 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
8941 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
9001 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
9061 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
9121 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
9181 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
9241 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
9301 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
9361 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
9421 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
9481 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
9541 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
9601 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
9661 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
9721 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
9781 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
9841 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
9901 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
9961 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
10021 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
10081 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
10141 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
10201 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
10261 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
10321 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
10381 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
10441 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
10501 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
10561 tccccgaaaa gtgccacctg ggtccttttc atcacgtgct ataaaaataa ttataattta
10621 aattttttaa tataaatata taaattaaaa atagaaagta aaaaaagaaa ttaaagaaaa
10681 aatagttttt gttttccgaa gatgtaaaag actctagggg gatcgccaac aaatactacc
10741 ttttatcttg ctcttcctgc tctcaggtat taatgccgaa ttgtttcatc ttgtctgtgt
10801 agaagaccac acacgaaaat cctgtgattt tacattttac ttatcgttaa tcgaatgtat
10861 atctatttaa tctgcttttc ttgtctaata aatatatatg taaagtacgc tttttgttga
10921 aattttttaa acctttgttt attttttttt cttcattccg taactcttct accttcttta
10981 tttactttct aaaatccaaa tacaaaacat aaaaataaat aaacacagag taaattccca
11041 aattattcca tcattaaaag atacgaggcg cgtgtaagtt acaggcaagc gatccgtcta
11101 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
11161 tc
//
