| 质粒名称: | pET-dCas9-VP64-6xHis |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 62935 |
| 存储实验室: | David Liu |
|
载体骨架基本信息 |
|
| 载体名称: | pET29 |
| 载体出品公司: | Novagen |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 5161 bp |
| 完整质粒大小: | 9544 bp |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 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: | dCas9-VP64 |
| 物种来源: | S. pyogenes |
| 插入大小: | 4383 bp |
| 基因突变: | D10A and H840A amino acid changes render Cas9 nuclease inactive for DNA cleavage |
| GenBank编号 | -- |
| 启动子 | T7 |
| 标签/融合蛋白: |
3xFLAG（插入件上的C端子）
NLS（插入件上的C端子）
VP64（插入件上的C端子）
6xHis（主干上的C终端）
|
| 克隆方法: | Gibson克隆 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | T7 promoter |
| 3' 测序引物 | T7 terminator |
|
文献引用方法 |
|
| 材料和方法部分: | pET-dCas9-VP64-6xHis was a gift from David Liu (Addgene plasmid # 62935 ; http://n2t.net/addgene:62935 ; RRID:Addgene_62935) |
| 参考文献部分: | Cationic lipid-mediated delivery of proteins enables efficient protein-based genome editing in vitro and in vivo. Zuris JA, Thompson DB, Shu Y, Guilinger JP, Bessen JL, Hu JH, Maeder ML, Joung JK, Chen ZY, Liu DR. Nat Biotechnol. 2015 Jan;33(1):73-80. doi: 10.1038/nbt.3081. Epub 2014 Oct 30. 10.1038/nbt.3081 PubMed 25357182 |
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 pET-dCas9-VP64-6xHis 9544 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Expression of dCas9-VP64-6xHis in bacterial cells.
ACCESSION .
VERSION .
KEYWORDS pET-dCas9-VP64-6xHis
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9544)
AUTHORS Zuris JA, Thompson DB, Shu Y, Guilinger JP, Bessen JL, Hu JH, Maeder
ML, Joung JK, Chen ZY, Liu DR
TITLE Cationic lipid-mediated delivery of proteins enables efficient
protein-based genome editing in vitro and in vivo.
JOURNAL Nat Biotechnol. 2015 Jan;33(1):73-80. doi: 10.1038/nbt.3081. Epub
2014 Oct 30.
PUBMED 25357182
REFERENCE 2 (bases 1 to 9544)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..9544
/organism="synthetic DNA construct"
/mol_type="other DNA"
protein_bind 3..27
/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)."
RBS 42..64
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 71..4174
/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="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
CDS 4181..4201
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 4208..4273
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS 4304..4453
/codon_start=1
/product="tetrameric repeat of the minimal activation
domain of herpes simplex virus VP16 (Beerli et al., 1998)"
/label=VP64
/translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
DLDML"
CDS 4454..4471
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 4490..4534
/codon_start=1
/product="affinity and epitope tag derived from pancreatic
ribonuclease A"
/label=S-Tag
/translation="KETAAAKFERQHMDS"
primer_bind complement(4572..4590)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 4586..4633
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 4670..5125
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4757..4776)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4967..4988
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(5240..6100)
/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 5863..5882
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(6101..6193)
/gene="bla"
/label=AmpR promoter
rep_origin 6274..6862
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6763..6782
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 7016..7033
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 7048..7190
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(7116..7135)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 7276..7298
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(7292..7483)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(8055..9137)
/codon_start=1
/gene="lacI"
/product="lac repressor"
/label=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)."
/translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
primer_bind 9099..9118
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(9138..9215)
/gene="lacI"
/label=lacI promoter
primer_bind 9421..9440
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 9528..3
/label=T7
/note="T7 promoter, forward primer"
promoter 9528..2
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
1 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
61 gagatatacc atggataaga aatactcaat aggcttagct atcggcacaa atagcgtcgg
121 atgggcggtg atcactgatg aatataaggt tccgtctaaa aagttcaagg ttctgggaaa
181 tacagaccgc cacagtatca aaaaaaatct tataggggct cttttatttg acagtggaga
241 gacagcggaa gcgactcgtc tcaaacggac agctcgtaga aggtatacac gtcggaagaa
301 tcgtatttgt tatctacagg agattttttc aaatgagatg gcgaaagtag atgatagttt
361 ctttcatcga cttgaagagt cttttttggt ggaagaagac aagaagcatg aacgtcatcc
421 tatttttgga aatatagtag atgaagttgc ttatcatgag aaatatccaa ctatctatca
481 tctgcgaaaa aaattggtag attctactga taaagcggat ttgcgcttaa tctatttggc
541 cttagcgcat atgattaagt ttcgtggtca ttttttgatt gagggagatt taaatcctga
601 taatagtgat gtggacaaac tatttatcca gttggtacaa acctacaatc aattatttga
661 agaaaaccct attaacgcaa gtggagtaga tgctaaagcg attctttctg cacgattgag
721 taaatcaaga cgattagaaa atctcattgc tcagctcccc ggtgagaaga aaaatggctt
781 atttgggaat ctcattgctt tgtcattggg tttgacccct aattttaaat caaattttga
841 tttggcagaa gatgctaaat tacagctttc aaaagatact tacgatgatg atttagataa
901 tttattggcg caaattggag atcaatatgc tgatttgttt ttggcagcta agaatttatc
961 agatgctatt ttactttcag atatcctaag agtaaatact gaaataacta aggctcccct
1021 atcagcttca atgattaaac gctacgatga acatcatcaa gacttgactc ttttaaaagc
1081 tttagttcga caacaacttc cagaaaagta taaagaaatc ttttttgatc aatcaaaaaa
1141 cggatatgca ggttatattg atgggggagc tagccaagaa gaattttata aatttatcaa
1201 accaatttta gaaaaaatgg atggtactga ggaattattg gtgaaactaa atcgtgaaga
1261 tttgctgcgc aagcaacgga cctttgacaa cggctctatt ccccatcaaa ttcacttggg
1321 tgagctgcat gctattttga gaagacaaga agacttttat ccatttttaa aagacaatcg
1381 tgagaagatt gaaaaaatct tgacttttcg aattccttat tatgttggtc cattggcgcg
1441 tggcaatagt cgttttgcat ggatgactcg gaagtctgaa gaaacaatta ccccatggaa
1501 ttttgaagaa gttgtcgata aaggtgcttc agctcaatca tttattgaac gcatgacaaa
1561 ctttgataaa aatcttccaa atgaaaaagt actaccaaaa catagtttgc tttatgagta
1621 ttttacggtt tataacgaat tgacaaaggt caaatatgtt actgaaggaa tgcgaaaacc
1681 agcatttctt tcaggtgaac agaagaaagc cattgttgat ttactcttca aaacaaatcg
1741 aaaagtaacc gttaagcaat taaaagaaga ttatttcaaa aaaatagaat gttttgatag
1801 tgttgaaatt tcaggagttg aagatagatt taatgcttca ttaggtacct accatgattt
1861 gctaaaaatt attaaagata aagatttttt ggataatgaa gaaaatgaag atatcttaga
1921 ggatattgtt ttaacattga ccttatttga agatagggag atgattgagg aaagacttaa
1981 aacatatgct cacctctttg atgataaggt gatgaaacag cttaaacgtc gccgttatac
2041 tggttgggga cgtttgtctc gaaaattgat taatggtatt agggataagc aatctggcaa
2101 aacaatatta gattttttga aatcagatgg ttttgccaat cgcaatttta tgcagctgat
2161 ccatgatgat agtttgacat ttaaagaaga cattcaaaaa gcacaagtgt ctggacaagg
2221 cgatagttta catgaacata ttgcaaattt agctggtagc cctgctatta aaaaaggtat
2281 tttacagact gtaaaagttg ttgatgaatt ggtcaaagta atggggcggc ataagccaga
2341 aaatatcgtt attgaaatgg cacgtgaaaa tcagacaact caaaagggcc agaaaaattc
2401 gcgagagcgt atgaaacgaa tcgaagaagg tatcaaagaa ttaggaagtc agattcttaa
2461 agagcatcct gttgaaaata ctcaattgca aaatgaaaag ctctatctct attatctcca
2521 aaatggaaga gacatgtatg tggaccaaga attagatatt aatcgtttaa gtgattatga
2581 tgtcgatgcc attgttccac aaagtttcct taaagacgat tcaatagaca ataaggtctt
2641 aacgcgttct gataaaaatc gtggtaaatc ggataacgtt ccaagtgaag aagtagtcaa
2701 aaagatgaaa aactattgga gacaacttct aaacgccaag ttaatcactc aacgtaagtt
2761 tgataattta acgaaagctg aacgtggagg tttgagtgaa cttgataaag ctggttttat
2821 caaacgccaa ttggttgaaa ctcgccaaat cactaagcat gtggcacaaa ttttggatag
2881 tcgcatgaat actaaatacg atgaaaatga taaacttatt cgagaggtta aagtgattac
2941 cttaaaatct aaattagttt ctgacttccg aaaagatttc caattctata aagtacgtga
3001 gattaacaat taccatcatg cccatgatgc gtatctaaat gccgtcgttg gaactgcttt
3061 gattaagaaa tatccaaaac ttgaatcgga gtttgtctat ggtgattata aagtttatga
3121 tgttcgtaaa atgattgcta agtctgagca agaaataggc aaagcaaccg caaaatattt
3181 cttttactct aatatcatga acttcttcaa aacagaaatt acacttgcaa atggagagat
3241 tcgcaaacgc cctctaatcg aaactaatgg ggaaactgga gaaattgtct gggataaagg
3301 gcgagatttt gccacagtgc gcaaagtatt gtccatgccc caagtcaata ttgtcaagaa
3361 aacagaagta cagacaggcg gattctccaa ggagtcaatt ttaccaaaaa gaaattcgga
3421 caagcttatt gctcgtaaaa aagactggga tccaaaaaaa tatggtggtt ttgatagtcc
3481 aacggtagct tattcagtcc tagtggttgc taaggtggaa aaagggaaat cgaagaagtt
3541 aaaatccgtt aaagagttac tagggatcac aattatggaa agaagttcct ttgaaaaaaa
3601 tccgattgac tttttagaag ctaaaggata taaggaagtt aaaaaagact taatcattaa
3661 actacctaaa tatagtcttt ttgagttaga aaacggtcgt aaacggatgc tggctagtgc
3721 cggagaatta caaaaaggaa atgagctggc tctgccaagc aaatatgtga attttttata
3781 tttagctagt cattatgaaa agttgaaggg tagtccagaa gataacgaac aaaaacaatt
3841 gtttgtggag cagcataagc attatttaga tgagattatt gagcaaatca gtgaattttc
3901 taagcgtgtt attttagcag atgccaattt agataaagtt cttagtgcat ataacaaaca
3961 tagagacaaa ccaatacgtg aacaagcaga aaatattatt catttattta cgttgacgaa
4021 tcttggagct cccgctgctt ttaaatattt tgatacaaca attgatcgta aacgatatac
4081 gtctacaaaa gaagttttag atgccactct tatccatcaa tccatcactg gtctttatga
4141 aacacgcatt gatttgagtc agctaggagg tgacggttct cccaagaaga agaggaaagt
4201 ctcgagcgac tacaaagacc atgacggtga ttataaagat catgacatcg attacaagga
4261 tgacgatgac aaggctgcag gaggcggtgg aagcgggcgc gccgacgcgc tggacgattt
4321 cgatctcgac atgctgggtt ctgatgccct cgatgacttt gacctggata tgttgggaag
4381 cgacgcattg gatgactttg atctggacat gctcggctcc gatgctctgg acgatttcga
4441 tctcgatatg ttacatcacc accaccatca ctaatgactc gagtctggta aagaaaccgc
4501 tgctgcgaaa tttgaacgcc agcacatgga ctcgtctact agcgcagctt aattaaccta
4561 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
4621 gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt
4681 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
4741 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
4801 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
4861 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
4921 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
4981 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
5041 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
5101 aacaaaatat taacgtttac aatttctggc ggcacgatgg catgagatta tcaaaaagga
5161 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
5221 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
5281 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
5341 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
5401 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
5461 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
5521 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
5581 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
5641 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
5701 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
5761 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
5821 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
5881 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
5941 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
6001 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
6061 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatcat
6121 gattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
6181 aaaaataaac aaataggtca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
6241 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
6301 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
6361 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
6421 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
6481 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
6541 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
6601 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
6661 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
6721 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
6781 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
6841 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
6901 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
6961 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
7021 gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg
7081 cggtatttca caccgcatat atggtgcact ctcagtacaa tctgctctga tgccgcatag
7141 ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc
7201 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
7261 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
7321 gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg
7381 cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga
7441 taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag
7501 ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac
7561 gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg
7621 tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata
7681 cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa
7741 tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca
7801 ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc
7861 gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca
7921 acgacaggag cacgatcatg ctagtcatgc cccgcgccca ccggaaggag ctgactgggt
7981 tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc taacttacat
8041 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
8101 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt
8161 cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt
8221 tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt
8281 aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatgtcc
8341 gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg
8401 ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga
8461 aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga
8521 gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc
8581 gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta
8641 ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat
8701 aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga
8761 tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag
8821 gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg
8881 cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca
8941 acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa
9001 ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc
9061 tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat
9121 aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc
9181 ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg
9241 cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc
9301 aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac
9361 catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc
9421 ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac
9481 gatgcgtccg gcgtagagga tcgagatcga tctcgatccc gcgaaattaa tacgactcac
9541 tata
//
