| 质粒名称: | pJSC129 - Bacterial expression plasmid for SpCas9-HF1, REC2 FRET variant |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 101211 |
| 存储实验室: | Jennifer Doudna, Keith Joung |
|
载体骨架基本信息 |
|
| 载体名称: | pCT10 |
| 载体出品公司: | Doudna lab |
| 空载体大小: | -- |
| 完整质粒大小: | 10073 bp |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 载体抗性: | 氨苄青霉素，100μg/mL |
| 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: | SpCas9 variant C80S/C574S/E60C/D273C/N497A/R661A/Q695A/Q926A |
| 物种来源: | Streptococcus pyogenes |
| 插入大小: | -- |
| 基因突变: | C80S, C574S, E60C, D273C, N497A, R661A, Q695A and Q926A |
| GenBank编号 | -- |
| 启动子: | T7 |
| 标签/融合蛋白: | 10xHis（主干上的N端子）MBP（主干上的N终端）TEV站点（主干上的N终端） |
| 克隆方法: | Gibson Cloning |
| 5’克隆位点: | -- |
| 3’克隆位点: | -- |
| 5’测序引物 | T7 |
| 3’测序引物 | T7-term |
|
文献引用方法 |
|
| 材料和方法部分: | pJSC129 - Bacterial expression plasmid for SpCas9-HF1, REC2 FRET variant was a gift from Jennifer Doudna & Keith Joung (Addgene plasmid # 101211 ; http://n2t.net/addgene:101211 ; RRID:Addgene_101211) |
| 参考文献部分: | Enhanced proofreading governs CRISPR-Cas9 targeting accuracy. Chen JS, Dagdas YS, Kleinstiver BP, Welch MM, Sousa AA, Harrington LB, Sternberg SH, Joung JK, Yildiz A, Doudna JA. Nature. 2017 Sep 20. doi: 10.1038/nature24268. 10.1038/nature24268 PubMed 28931002 |
pJSC129 - Bacterial expression plasmid for SpCas9-HF1, REC2
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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 pJSC129 - Bacterial expression plasmid for SpCas9-HF1, REC2 FRET variant 10073 bp ds-DNA circular SYN 13-MAY-2021
DEFINITION Bacterial expression plasmid for SpCas9-HF1, REC2 FRET variant.
ACCESSION .
VERSION .
KEYWORDS pJSC129 - Bacterial expression plasmid for SpCas9-HF1, REC2 FR...
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 10073)
AUTHORS Chen JS, Dagdas YS, Kleinstiver BP, Welch MM, Sousa AA, Harrington
LB, Sternberg SH, Joung JK, Yildiz A, Doudna JA
TITLE Enhanced proofreading governs CRISPR-Cas9 targeting accuracy.
JOURNAL Nature. 2017 Sep 20. doi: 10.1038/nature24268.
PUBMED 28931002
REFERENCE 2 (bases 1 to 10073)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..10073
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 120..139
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 223..242
/label=T7
/note="T7 promoter, forward primer"
promoter 223..241
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
RBS 330..352
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 372..401
/codon_start=1
/product="10xHis affinity tag"
/label=10xHis
/translation="HHHHHHHHHH"
CDS 411..1511
/codon_start=1
/gene="malE (mutated)"
/product="maltose binding protein from E. coli"
/label=MBP
/note="This version of the gene does not encode a signal
sequence, so MBP will remain in the cytosol."
/translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
WYAVRTAVINAASGRQTVDEALKDAQT"
primer_bind 1485..1508
/label=MBP-F
/note="Maltose binding protein, forward primer"
CDS 1566..1586
/codon_start=1
/product="tobacco etch virus (TEV) protease recognition and
cleavage site"
/label=TEV site
/translation="ENLYFQS"
CDS 1593..5699
/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
NLIGALLFDSGETACATRLKRTARRRYTRRKNRISYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDCDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTAFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIESFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGALSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
ALIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRAITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
primer_bind complement(5825..5843)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 5839..5886
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
promoter complement(6252..6280)
/label=tet promoter
/note="E. coli promoter for tetracycline efflux protein
gene"
primer_bind complement(6307..6325)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 6393..6497
/gene="bla"
/label=AmpR promoter
CDS 6498..7358
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(6716..6735)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 7529..8117
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 8018..8037
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 8271..8288
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 8303..8442
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(8368..8387)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 8528..8550
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(8544..8735)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
ORIGIN
1 aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc
61 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
121 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
181 atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga ctcactatag
241 ggagaccaca acggtttccc tctagtgccg gctccggaga gctctttaat taagcggccg
301 ccctgcagga ctcgagttct agaaataatt ttgtttaact ttaagaagga gatatacata
361 tgaaatcttc tcaccatcac catcaccatc accatcacca tggttcttct atgaaaatcg
421 aagaaggtaa actggtaatc tggattaacg gcgataaagg ctataacggt ctcgctgaag
481 tcggtaagaa attcgagaaa gataccggaa ttaaagtcac cgttgagcat ccggataaac
541 tggaagagaa attcccacag gttgcggcaa ctggcgatgg ccctgacatt atcttctggg
601 cacacgaccg ctttggtggc tacgctcaat ctggcctgtt ggctgaaatc accccggaca
661 aagcgttcca ggacaagctg tatccgttta cctgggatgc cgtacgttac aacggcaagc
721 tgattgctta cccgatcgct gttgaagcgt tatcgctgat ttataacaaa gatctgctgc
781 cgaacccgcc aaaaacctgg gaagagatcc cggcgctgga taaagaactg aaagcgaaag
841 gtaagagcgc gctgatgttc aacctgcaag aaccgtactt cacctggccg ctgattgctg
901 ctgacggggg ttatgcgttc aagtatgaaa acggcaagta cgacattaaa gacgtgggcg
961 tggataacgc tggcgcaaaa gcgggtctga ccttcctggt tgacctgatt aaaaacaaac
1021 acatgaatgc agacaccgat tactccatcg cagaagctgc ctttaataaa ggcgaaacag
1081 cgatgaccat caacggcccg tgggcatggt ccaacatcga caccagcaaa gtgaattatg
1141 gtgtaacggt actgccgacc ttcaagggtc aaccatccaa accgttcgtt ggcgtgctga
1201 gcgcaggtat taacgccgcc agtccgaaca aagagctggc aaaagagttc ctcgaaaact
1261 atctgctgac tgatgaaggt ctggaagcgg ttaataaaga caaaccgctg ggtgccgtag
1321 cgctgaagtc ttacgaggaa gagttggcga aagatccacg tattgccgcc actatggaaa
1381 acgcccagaa aggtgaaatc atgccgaaca tcccgcagat gtccgctttc tggtatgccg
1441 tgcgtactgc ggtgatcaac gccgccagcg gtcgtcagac tgtcgatgaa gccctgaaag
1501 acgcgcagac taattcgagc tcgaacaaca acaacaataa caataacaac aacctcggga
1561 tcgaggaaaa cctgtacttc caatccaatg caatggataa gaaatactca ataggcttag
1621 atatcggcac aaatagcgtc ggatgggcgg tgatcactga tgaatataag gttccgtcta
1681 aaaagttcaa ggttctggga aatacagacc gccacagtat caaaaaaaat cttatagggg
1741 ctcttttatt tgacagtgga gagacagcgt gtgcgactcg tctcaaacgg acagctcgta
1801 gaaggtatac acgtcggaag aatcgtattt cttatctaca ggagattttt tcaaatgaga
1861 tggcgaaagt agatgatagt ttctttcatc gacttgaaga gtcttttttg gtggaagaag
1921 acaagaagca tgaacgtcat cctatttttg gaaatatagt agatgaagtt gcttatcatg
1981 agaaatatcc aactatctat catctgcgaa aaaaattggt agattctact gataaagcgg
2041 atttgcgctt aatctatttg gccttagcgc atatgattaa gtttcgtggt cattttttga
2101 ttgagggaga tttaaatcct gataatagtg atgtggacaa actatttatc cagttggtac
2161 aaacctacaa tcaattattt gaagaaaacc ctattaacgc aagtggagta gatgctaaag
2221 cgattctttc tgcacgattg agtaaatcaa gacgattaga aaatctcatt gctcagctcc
2281 ccggtgagaa gaaaaatggc ttatttggga atctcattgc tttgtcattg ggtttgaccc
2341 ctaattttaa atcaaatttt gatttggcag aagatgctaa attacagctt tcaaaagata
2401 cttacgattg tgatttagat aatttattgg cgcaaattgg agatcaatat gctgatttgt
2461 ttttggcagc taagaattta tcagatgcta ttttactttc agatatccta agagtaaata
2521 ctgaaataac taaggctccc ctatcagctt caatgattaa acgctacgat gaacatcatc
2581 aagacttgac tcttttaaaa gctttagttc gacaacaact tccagaaaag tataaagaaa
2641 tcttttttga tcaatcaaaa aacggatatg caggttatat tgatggggga gctagccaag
2701 aagaatttta taaatttatc aaaccaattt tagaaaaaat ggatggtact gaggaattat
2761 tggtgaaact aaatcgtgaa gatttgctgc gcaagcaacg gacctttgac aacggctcta
2821 ttccccatca aattcacttg ggtgagctgc atgctatttt gagaagacaa gaagactttt
2881 atccattttt aaaagacaat cgtgagaaga ttgaaaaaat cttgactttt cgaattcctt
2941 attatgttgg tccattggcg cgtggcaata gtcgttttgc atggatgact cggaagtctg
3001 aagaaacaat taccccatgg aattttgaag aagttgtcga taaaggtgct tcagctcaat
3061 catttattga acgcatgaca gcgtttgata aaaatcttcc aaatgaaaaa gtactaccaa
3121 aacatagttt gctttatgag tattttacgg tttataacga attgacaaag gtcaaatatg
3181 ttactgaagg aatgcgaaaa ccagcatttc tttcaggtga acagaagaaa gccattgttg
3241 atttactctt caaaacaaat cgaaaagtaa ccgttaagca attaaaagaa gattatttca
3301 aaaaaataga atcttttgat agtgttgaaa tttcaggagt tgaagataga tttaatgctt
3361 cattaggtac ctaccatgat ttgctaaaaa ttattaaaga taaagatttt ttggataatg
3421 aagaaaatga agatatctta gaggatattg ttttaacatt gaccttattt gaagataggg
3481 agatgattga ggaaagactt aaaacatatg ctcacctctt tgatgataag gtgatgaaac
3541 agcttaaacg tcgccgttat actggttggg gagcgttgtc tcgaaaattg attaatggta
3601 ttagggataa gcaatctggc aaaacaatat tagatttttt gaaatcagat ggttttgcca
3661 atcgcaattt tatggccctg atccatgatg atagtttgac atttaaagaa gacattcaaa
3721 aagcacaagt gtctggacaa ggcgatagtt tacatgaaca tattgcaaat ttagctggta
3781 gccctgctat taaaaaaggt attttacaga ctgtaaaagt tgttgatgaa ttggtcaaag
3841 taatggggcg gcataagcca gaaaatatcg ttattgaaat ggcacgtgaa aatcagacaa
3901 ctcaaaaggg ccagaaaaat tcgcgagagc gtatgaaacg aatcgaagaa ggtatcaaag
3961 aattaggaag tcagattctt aaagagcatc ctgttgaaaa tactcaattg caaaatgaaa
4021 agctctatct ctattatctc caaaatggaa gagacatgta tgtggaccaa gaattagata
4081 ttaatcgttt aagtgattat gatgtcgatc acattgttcc acaaagtttc cttaaagacg
4141 attcaataga caataaggtc ttaacgcgtt ctgataaaaa tcgtggtaaa tcggataacg
4201 ttccaagtga agaagtagtc aaaaagatga aaaactattg gagacaactt ctaaacgcca
4261 agttaatcac tcaacgtaag tttgataatt taacgaaagc tgaacgtgga ggtttgagtg
4321 aacttgataa agctggtttt atcaaacgcc aattggttga aactcgcgcc atcactaagc
4381 atgtggcaca aattttggat agtcgcatga atactaaata cgatgaaaat gataaactta
4441 ttcgagaggt taaagtgatt accttaaaat ctaaattagt ttctgacttc cgaaaagatt
4501 tccaattcta taaagtacgt gagattaaca attaccatca tgcccatgat gcgtatctaa
4561 atgccgtcgt tggaactgct ttgattaaga aatatccaaa acttgaatcg gagtttgtct
4621 atggtgatta taaagtttat gatgttcgta aaatgattgc taagtctgag caagaaatag
4681 gcaaagcaac cgcaaaatat ttcttttact ctaatatcat gaacttcttc aaaacagaaa
4741 ttacacttgc aaatggagag attcgcaaac gccctctaat cgaaactaat ggggaaactg
4801 gagaaattgt ctgggataaa gggcgagatt ttgccacagt gcgcaaagta ttgtccatgc
4861 cccaagtcaa tattgtcaag aaaacagaag tacagacagg cggattctcc aaggagtcaa
4921 ttttaccaaa aagaaattcg gacaagctta ttgctcgtaa aaaagactgg gatccaaaaa
4981 aatatggtgg ttttgatagt ccaacggtag cttattcagt cctagtggtt gctaaggtgg
5041 aaaaagggaa atcgaagaag ttaaaatccg ttaaagagtt actagggatc acaattatgg
5101 aaagaagttc ctttgaaaaa aatccgattg actttttaga agctaaagga tataaggaag
5161 ttaaaaaaga cttaatcatt aaactaccta aatatagtct ttttgagtta gaaaacggtc
5221 gtaaacggat gctggctagt gccggagaat tacaaaaagg aaatgagctg gctctgccaa
5281 gcaaatatgt gaatttttta tatttagcta gtcattatga aaagttgaag ggtagtccag
5341 aagataacga acaaaaacaa ttgtttgtgg agcagcataa gcattattta gatgagatta
5401 ttgagcaaat cagtgaattt tctaagcgtg ttattttagc agatgccaat ttagataaag
5461 ttcttagtgc atataacaaa catagagaca aaccaatacg tgaacaagca gaaaatatta
5521 ttcatttatt tacgttgacg aatcttggag ctcccgctgc ttttaaatat tttgatacaa
5581 caattgatcg taaacgatat acgtctacaa aagaagtttt agatgccact cttatccatc
5641 aatccatcac tggtctttat gaaacacgca ttgatttgag tcagctagga ggtgactaat
5701 aacattggaa gtggataacg gatccgcgat cgcggcgcgc cacctggtgg ccggccggta
5761 ccacgcgtgc gcgctgatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct
5821 gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt
5881 tttttgctga aaggaggaac tatatccgga tatccacagg acgggtgtgg tcgccatgat
5941 cgcgtagtcg atagtggctc caagtagcga agcgagcagg actgggcggc ggccaaagcg
6001 gtcggacagt gctccgagaa cgggtgcgca tagaaattgc atcaacgcat atagcgctag
6061 cagcacgcca tagtgactgg cgatgctgtc ggaatggacg atatcccgca agaggcccgg
6121 cagtaccggc ataaccaagc ctatgcctac agcatccagg gtgacggtgc cgaggatgac
6181 gatgagcgca ttgttagatt tcatacacgg tgcctgactg cgttagcaat ttaactgtga
6241 taaactaccg cattaaagct tatcgatgat aagctgtcaa acatgagaat tcttgaagac
6301 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
6361 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
6421 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataac
6481 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
6541 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
6601 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
6661 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
6721 gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc cgcatacact
6781 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
6841 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
6901 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
6961 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
7021 agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg
7081 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
7141 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
7201 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
7261 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
7321 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
7381 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
7441 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
7501 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
7561 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
7621 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
7681 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
7741 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
7801 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
7861 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
7921 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
7981 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
8041 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
8101 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
8161 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
8221 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
8281 tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
8341 tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
8401 gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca
8461 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
8521 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg
8581 cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca
8641 tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg
8701 gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt
8761 ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact
8821 gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg
8881 cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta
8941 ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag
9001 ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt
9061 gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt
9121 gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg
9181 agcacgatca tgcgcacccg tggccaggac ccaacgctgc ccgagatgcg ccgcgtgcgg
9241 ctgctggaga tggcggacgc gatggatatg ttctgccaag ggttggtttg cgcattcaca
9301 gttctccgca agaattgatt ggctccaatt cttggagtgg tgaatccgtt agcgaggtgc
9361 cgccggcttc cattcaggtc gaggtggccc ggctccatgc accgcgacgc aacgcgggga
9421 ggcagacaag gtatagggcg gcgcctacaa tccatgccaa cccgttccat gtgctcgccg
9481 aggcggcata aatcgccgtg acgatcagcg gtccaatgat cgaagttagg ctggtaagag
9541 ccgcgagcga tccttgaagc tgtccctgat ggtcgtcatc tacctgcctg gacagcatgg
9601 cctgcaacgc gggcatcccg atgccgccgg aagcgagaag aatcataatg gggaaggcca
9661 tccagcctcg cgtcgcgaac gccagcaaga cgtagcccag cgcgtcggcc gccatgccgg
9721 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
9781 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
9841 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
9901 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
9961 agctgactgg gttgaaggct ctcaagggca tcggtcgacg ctctccctta tgcgactcct
10021 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gca
//
