| 质粒名称: | pREDCas9 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 71541 |
| 存储实验室: | Tao Chen |
|
载体骨架基本信息 |
|
| 载体名称: | pSC101ts |
| 载体出品公司: | -- |
| 载体抗性: | 大观霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 低拷贝 |
| 生长条件: | 30 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | Cas9 |
| 物种来源: | S. pyogenes |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | -- |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | -- |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pREDCas9 was a gift from Tao Chen (Addgene plasmid # 71541 ; http://n2t.net/addgene:71541 ; RRID:Addgene_71541) |
| 参考文献部分: | Metabolic engineering of Escherichia coli using CRISPR-Cas9 meditated genome editing. Li Y, Lin Z, Huang C, Zhang Y, Wang Z, Tang YJ, Chen T, Zhao X. Metab Eng. 2015 Sep;31:13-21. doi: 10.1016/j.ymben.2015.06.006. Epub 2015 Jun 30. 10.1016/j.ymben.2015.06.006 PubMed 26141150 |
买家导航
pREDCas9载体质粒基本信息
pREDCas9质粒图谱载体图谱和pREDCas9载体序列质粒序列多克隆位点信息
pREDCas9质粒载体简介
pREDCas9质粒序列载体序列
LOCUS pREDCas9 15460 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Constitutive expression of Cas9, inducible expression of the Red
recombineering system, and inducible expression of the plasmid
curing system for CRISPR mediated genome editing of E. coli..
ACCESSION .
VERSION .
KEYWORDS pREDCas9
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 15460)
AUTHORS Li Y, Lin Z, Huang C, Zhang Y, Wang Z, Tang YJ, Chen T, Zhao X
TITLE Metabolic engineering of Escherichia coli using CRISPR-Cas9
meditated genome editing.
JOURNAL Metab Eng. 2015 Sep;31:13-21. doi: 10.1016/j.ymben.2015.06.006. Epub
2015 Jun 30.
PUBMED 26141150
REFERENCE 2 (bases 1 to 15460)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..15460
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_RNA complement(370..448)
/label=tracrRNA
/note="trans-activating CRISPR RNA for the Streptococcus
pyogenes CRISPR/Cas9 system"
CDS 744..4850
/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"
misc_feature 4871..5002
/label=crRNA leader
/note="crRNA leader sequence for the Streptococcus pyogenes
CRISPR/Cas system"
repeat_region 5003..5038
/label=DR
/note="direct repeat for the Streptococcus pyogenes
CRISPR/Cas system"
CDS complement(5248..6126)
/codon_start=1
/gene="araC"
/product="L-arabinose regulatory protein"
/label=araC
/translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
EKVNDVAVKLS"
promoter 6153..6437
/gene="araBAD"
/label=araBAD promoter
/note="promoter of the L-arabinose operon of E. coli; the
araC regulatory gene is transcribed in the opposite
direction (Guzman et al., 1995)"
primer_bind 6360..6379
/label=pBAD Forward
/note="For vectors with E. coli araBAD promoter, forward
primer"
repeat_region 6584..6619
/label=DR
/note="direct repeat for the Streptococcus pyogenes
CRISPR/Cas system"
CDS 7397..8188
/codon_start=1
/gene="aadA"
/product="aminoglycoside adenylyltransferase (Murphy,
1985)"
/label=SmR
/note="confers resistance to spectinomycin and
streptomycin"
/translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAAYWAMERLPAQYQP
VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
CDS complement(8512..9192)
/codon_start=1
/gene="exo"
/product="5' to 3' double-stranded DNA exonuclease in the
lambda Red system"
/label=Exo
/note="creates 3' single-stranded DNA tails"
/translation="MTPDIILQRTGIDVRAVEQGEDAWHKLRLGVITASEVHNVIAKPR
SGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESP
IIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMW
VTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFG
EQWR"
CDS complement(9189..9974)
/codon_start=1
/gene="bet"
/product="single-stranded DNA binding recombinase in the
lambda Red system"
/label=Beta
/note="mediates pairing of a single-stranded DNA with a
complementary target"
/translation="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFI
ALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESC
TCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLA
FGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLP
LCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA"
CDS complement(9980..10396)
/codon_start=1
/gene="gam"
/product="inhibitor of the host RecBCD nuclease in the
lambda Red system"
/label=Gam
/note="protects the double-stranded DNA substrate for
recombination"
/translation="MDINTETEIKQKHSLTPFPVFLISPAFRGRYFHSYFRSSAMNAYY
IQDRLEAQSWARHYQQLAREEKEAELADDMEKGLPQHLFESLCIDHLQRHGASKKSITR
AFDDDVEFQERMAEHIRYMVETIAHHQVDIDSEV"
primer_bind complement(10419..10441)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 10430..10446
/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(10454..10484)
/label=lac UV5 promoter
/note="E. coli lac promoter with an ""up"" mutation"
CDS complement(10610..11671)
/codon_start=1
/gene="recA"
/product="DNA repair protein from E. coli"
/label=RecA
/translation="MAIDENKQKALAAALGQIEKQFGKGSIMRLGEDRSMDVETISTGS
LSLDIALGAGGLPMGRIVEIYGPESSGKTTLTLQVIAAAQREGKTCAFIDAEHALDPIY
ARKLGVDIDNLLCSQPDTGEQALEICDALARSGAVDVIVVDSVAALTPKAEIEGEIGDS
HMGLAARMMSQAMRKLAGNLKQSNTLLIFINQIRMKIGVMFGNPETTTGGNALKFYASV
RLDIRRIGAVKEGENVVGSETRVKVVKNKIAAPFKQAEFQILYGEGINFYGELVDLGVK
EKLIEKAGAWYSYKGEKIGQGKANATAWLKDNPETAKEIEKKVRELLLSNPNSTPDFSV
DDSEGVAETNEDF"
CDS complement(12160..13242)
/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 13204..13223
/label=LacI-R
/note="LacI, reverse primer"
CDS complement(13428..14378)
/codon_start=1
/gene="rep101"
/product="RepA protein needed for replication with the
pSC101 origin"
/label=Rep101
/translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
rep_origin 14426..14648
/label=pSC101 ori
/note="low-copy replication origin that requires the Rep101
protein"
ORIGIN
1 ctgctcgcgc aggctgggtg ccaagctctc gggtaacatc aaggcccgat ccttggagcc
61 cttgccctcc cgcacgatga tcgtgccgtg atcgaaatcc agatccttga cccgcagttg
121 caaaccctca ctgatccgca tgcggatctt gctgtaggca taggcttggt tatgccggta
181 ctgccgggcc tcttgcggga ttacgaaatc atcctgtgga gcttagtagg tttagcaaga
241 tggcagcgcc taaatgtaga atgataaaag gattaagaga ttaatttccc taaaaatgat
301 aaaacaagcg ttttgaaagc gcttgttttt ttggtttgca gtcagagtag aatagaagta
361 tcaaaaaaag caccgactcg gtgccacttt ttcaagttga taacggacta gccttatttt
421 aacttgctat gctgttttga atggttccaa caagattatt ttataacttt tataacaaat
481 aatcaaggag aaattcaaag aaatttatca gccataaaac aatacttaat actatagaat
541 gataacaaaa taaactactt tttaaaagaa ttttgtgtta taatctattt attattaagt
601 attgggtaat attttttgaa gagatatttt gaaaaagaaa aattaaagca tattaaacta
661 atttcggagg tcattaaaac tattattgaa atcatcaaac tcattatgga tttaatttaa
721 actttttatt ttaggaggca aaaatggata agaaatactc aataggctta gatatcggca
781 caaatagcgt cggatgggcg gtgatcactg atgaatataa ggttccgtct aaaaagttca
841 aggttctggg aaatacagac cgccacagta tcaaaaaaaa tcttataggg gctcttttat
901 ttgacagtgg agagacagcg gaagcgactc gtctcaaacg gacagctcgt agaaggtata
961 cacgtcggaa gaatcgtatt tgttatctac aggagatttt ttcaaatgag atggcgaaag
1021 tagatgatag tttctttcat cgacttgaag agtctttttt ggtggaagaa gacaagaagc
1081 atgaacgtca tcctattttt ggaaatatag tagatgaagt tgcttatcat gagaaatatc
1141 caactatcta tcatctgcga aaaaaattgg tagattctac tgataaagcg gatttgcgct
1201 taatctattt ggccttagcg catatgatta agtttcgtgg tcattttttg attgagggag
1261 atttaaatcc tgataatagt gatgtggaca aactatttat ccagttggta caaacctaca
1321 atcaattatt tgaagaaaac cctattaacg caagtggagt agatgctaaa gcgattcttt
1381 ctgcacgatt gagtaaatca agacgattag aaaatctcat tgctcagctc cccggtgaga
1441 agaaaaatgg cttatttggg aatctcattg ctttgtcatt gggtttgacc cctaatttta
1501 aatcaaattt tgatttggca gaagatgcta aattacagct ttcaaaagat acttacgatg
1561 atgatttaga taatttattg gcgcaaattg gagatcaata tgctgatttg tttttggcag
1621 ctaagaattt atcagatgct attttacttt cagatatcct aagagtaaat actgaaataa
1681 ctaaggctcc cctatcagct tcaatgatta aacgctacga tgaacatcat caagacttga
1741 ctcttttaaa agctttagtt cgacaacaac ttccagaaaa gtataaagaa atcttttttg
1801 atcaatcaaa aaacggatat gcaggttata ttgatggggg agctagccaa gaagaatttt
1861 ataaatttat caaaccaatt ttagaaaaaa tggatggtac tgaggaatta ttggtgaaac
1921 taaatcgtga agatttgctg cgcaagcaac ggacctttga caacggctct attccccatc
1981 aaattcactt gggtgagctg catgctattt tgagaagaca agaagacttt tatccatttt
2041 taaaagacaa tcgtgagaag attgaaaaaa tcttgacttt tcgaattcct tattatgttg
2101 gtccattggc gcgtggcaat agtcgttttg catggatgac tcggaagtct gaagaaacaa
2161 ttaccccatg gaattttgaa gaagttgtcg ataaaggtgc ttcagctcaa tcatttattg
2221 aacgcatgac aaactttgat aaaaatcttc caaatgaaaa agtactacca aaacatagtt
2281 tgctttatga gtattttacg gtttataacg aattgacaaa ggtcaaatat gttactgaag
2341 gaatgcgaaa accagcattt ctttcaggtg aacagaagaa agccattgtt gatttactct
2401 tcaaaacaaa tcgaaaagta accgttaagc aattaaaaga agattatttc aaaaaaatag
2461 aatgttttga tagtgttgaa atttcaggag ttgaagatag atttaatgct tcattaggta
2521 cctaccatga tttgctaaaa attattaaag ataaagattt tttggataat gaagaaaatg
2581 aagatatctt agaggatatt gttttaacat tgaccttatt tgaagatagg gagatgattg
2641 aggaaagact taaaacatat gctcacctct ttgatgataa ggtgatgaaa cagcttaaac
2701 gtcgccgtta tactggttgg ggacgtttgt ctcgaaaatt gattaatggt attagggata
2761 agcaatctgg caaaacaata ttagattttt tgaaatcaga tggttttgcc aatcgcaatt
2821 ttatgcagct gatccatgat gatagtttga catttaaaga agacattcaa aaagcacaag
2881 tgtctggaca aggcgatagt ttacatgaac atattgcaaa tttagctggt agccctgcta
2941 ttaaaaaagg tattttacag actgtaaaag ttgttgatga attggtcaaa gtaatggggc
3001 ggcataagcc agaaaatatc gttattgaaa tggcacgtga aaatcagaca actcaaaagg
3061 gccagaaaaa ttcgcgagag cgtatgaaac gaatcgaaga aggtatcaaa gaattaggaa
3121 gtcagattct taaagagcat cctgttgaaa atactcaatt gcaaaatgaa aagctctatc
3181 tctattatct ccaaaatgga agagacatgt atgtggacca agaattagat attaatcgtt
3241 taagtgatta tgatgtcgat cacattgttc cacaaagttt ccttaaagac gattcaatag
3301 acaataaggt cttaacgcgt tctgataaaa atcgtggtaa atcggataac gttccaagtg
3361 aagaagtagt caaaaagatg aaaaactatt ggagacaact tctaaacgcc aagttaatca
3421 ctcaacgtaa gtttgataat ttaacgaaag ctgaacgtgg aggtttgagt gaacttgata
3481 aagctggttt tatcaaacgc caattggttg aaactcgcca aatcactaag catgtggcac
3541 aaattttgga tagtcgcatg aatactaaat acgatgaaaa tgataaactt attcgagagg
3601 ttaaagtgat taccttaaaa tctaaattag tttctgactt ccgaaaagat ttccaattct
3661 ataaagtacg tgagattaac aattaccatc atgcccatga tgcgtatcta aatgccgtcg
3721 ttggaactgc tttgattaag aaatatccaa aacttgaatc ggagtttgtc tatggtgatt
3781 ataaagttta tgatgttcgt aaaatgattg ctaagtctga gcaagaaata ggcaaagcaa
3841 ccgcaaaata tttcttttac tctaatatca tgaacttctt caaaacagaa attacacttg
3901 caaatggaga gattcgcaaa cgccctctaa tcgaaactaa tggggaaact ggagaaattg
3961 tctgggataa agggcgagat tttgccacag tgcgcaaagt attgtccatg ccccaagtca
4021 atattgtcaa gaaaacagaa gtacagacag gcggattctc caaggagtca attttaccaa
4081 aaagaaattc ggacaagctt attgctcgta aaaaagactg ggatccaaaa aaatatggtg
4141 gttttgatag tccaacggta gcttattcag tcctagtggt tgctaaggtg gaaaaaggga
4201 aatcgaagaa gttaaaatcc gttaaagagt tactagggat cacaattatg gaaagaagtt
4261 cctttgaaaa aaatccgatt gactttttag aagctaaagg atataaggaa gttaaaaaag
4321 acttaatcat taaactacct aaatatagtc tttttgagtt agaaaacggt cgtaaacgga
4381 tgctggctag tgccggagaa ttacaaaaag gaaatgagct ggctctgcca agcaaatatg
4441 tgaatttttt atatttagct agtcattatg aaaagttgaa gggtagtcca gaagataacg
4501 aacaaaaaca attgtttgtg gagcagcata agcattattt agatgagatt attgagcaaa
4561 tcagtgaatt ttctaagcgt gttattttag cagatgccaa tttagataaa gttcttagtg
4621 catataacaa acatagagac aaaccaatac gtgaacaagc agaaaatatt attcatttat
4681 ttacgttgac gaatcttgga gctcccgctg cttttaaata ttttgataca acaattgatc
4741 gtaaacgata tacgtctaca aaagaagttt tagatgccac tcttatccat caatccatca
4801 ctggtcttta tgaaacacgc attgatttga gtcagctagg aggtgactga agtatatttt
4861 agatgaagat tatttcttaa taactaaaaa tatggtataa tactcttaat aaatgcagta
4921 atacaggggc ttttcaagac tgaagtctag ctgagacaaa tagtgcgatt acgaaatttt
4981 ttagacaaaa atagtctacg aggttttaga gctatgctgt tttgaatggt cccaaaactg
5041 cagcgcaata gttggcgaag taatcgcaac atccgcatta aaatctagcg agggctttac
5101 taagctccgc aaaaaacccc gcccctgaca gggcggggtt ttttcgcgcc cgatccttgg
5161 agcccttgcc ctcccgcacg atgatcgtgc cgtgatcgaa atccagatcc ttgacccgca
5221 gttgcaaacc ctcactgatc cgcatgctta tgacaacttg acggctacat cattcacttt
5281 ttcttcacaa ccggcacgga actcgctcgg gctggccccg gtgcattttt taaatacccg
5341 cgagaaatag agttgatcgt caaaaccaac attgcgaccg acggtggcga taggcatccg
5401 ggtggtgctc aaaagcagct tcgcctggct gatacgttgg tcctcgcgcc agcttaagac
5461 gctaatccct aactgctggc ggaaaagatg tgacagacgc gacggcgaca agcaaacatg
5521 ctgtgcgacg ctggcgatat caaaattgct gtctgccagg tgatcgctga tgtactgaca
5581 agcctcgcgt acccgattat ccatcggtgg atggagcgac tcgttaatcg cttccatgcg
5641 ccgcagtaac aattgctcaa gcagatttat cgccagcagc tccgaatagc gcccttcccc
5701 ttgcccggcg ttaatgattt gcccaaacag gtcgctgaaa tgcggctggt gcgcttcatc
5761 cgggcgaaag aaccccgtat tggcaaatat tgacggccag ttaagccatt catgccagta
5821 ggcgcgcgga cgaaagtaaa cccactggtg ataccattcg cgagcctccg gatgacgacc
5881 gtagtgatga atctctcctg gcgggaacag caaaatatca cccggtcggc aaacaaattc
5941 tcgtccctga tttttcacca ccccctgacc gcgaatggtg agattgagaa tataaccttt
6001 cattcccagc ggtcggtcga taaaaaaatc gagataaccg ttggcctcaa tcggcgttaa
6061 acccgccacc agatgggcat taaacgagta tcccggcagc aggggatcat tttgcgcttc
6121 agccatactt ttcatactcc cgccattcag agaagaaacc aattgtccat attgcatcag
6181 acattgccgt cactgcgtct tttactggct cttctcgcta accaaaccgg taaccccgct
6241 tattaaaagc attctgtaac aaagcgggac caaagccatg acaaaaacgc gtaacaaaag
6301 tgtctataat cacggcagaa aagtccacat tgattatttg cacggcgtca cactttgcta
6361 tgccatagca tttttatcca taagattagc ggatcctacc tgacgctttt tatcgcaact
6421 ctctactgtt tctccatgta ggtcgttcgc tccaagcgtt ttagagctag aaatagcaag
6481 ttaaaataag gctagtccgt tatcaacttg aaaaagtggc accagacccc gttgatgata
6541 ccgctgcctt actgggtgca ttagccagtc tgaatgacct cgagttttag agctatgctg
6601 ttttgaatgg tcccaaaact tcagcacact gagacttgtt gagttccatg ttttagagct
6661 atgctgtttt gaatggactc cattcaacat tgccgatgat aacttgagaa agagggttaa
6721 taccagcagt cggatacctt cctattcttt ctgttaaagc gttttcatgt tataataggc
6781 aaaagaagag tagtgtgatc gtccattccg acgtcgagcc gttccataca gaagctgggc
6841 gaacaaacga tgctcgcctt ccagaaaacc gaggatgcga accacttcat ccggggtcag
6901 caccaccggc aagcgccgcg acggccgagg tcttccgatc tcctgaagcc agggcagatc
6961 cgtgcacagc accttgccgt agaagaacag caaggccgcc aatgcctgac gatgcgtgga
7021 gaccgaaacc ttgcgctcgt tcgccagcca ggacagaaat gcctcgactt cgctgctgcc
7081 caaggttgcc gggtgacgca caccgtggaa acggatgaag gcacgaaccc agtggacata
7141 agcctgttcg gttcgtaagc tgtaatgcaa gtagcgtatg cgctcacgca actggtccag
7201 aaccttgacc gaacgcagcg gtggtaacgg cgcagtggcg gttttcatgg cttgttatga
7261 ctgttttttt ggggtacagt ctatgcctcg ggcatccaag cagcaagcgc gttacgccgt
7321 gggtcgatgt ttgatgttat ggagcagcaa cgatgttacg cagcagggca gtcgccctaa
7381 aacaaagtta aacatcatga gggaagcggt gatcgccgaa gtatcgactc aactatcaga
7441 ggtagttggc gtcatcgagc gccatctcga accgacgttg ctggccgtac atttgtacgg
7501 ctccgcagtg gatggcggcc tgaagccaca cagtgatatt gatttgctgg ttacggtgac
7561 cgtaaggctt gatgaaacaa cgcggcgagc tttgatcaac gaccttttgg aaacttcggc
7621 ttcccctgga gagagcgaga ttctccgcgc tgtagaagtc accattgttg tgcacgacga
7681 catcattccg tggcgttatc cagctaagcg cgaactgcaa tttggagaat ggcagcgcaa
7741 tgacattctt gcaggtatct tcgagccagc cacgatcgac attgatctgg ctatcttgct
7801 gacaaaagca agagaacata gcgttgcctt ggtaggtcca gcggcggagg aactctttga
7861 tccggttcct gaacaggatc tatttgaggc gctaaatgaa accttaacgc tatggaactc
7921 gccgcccgac tgggctggcg atgagcgaaa tgtagtgctt acgttgtccc gcatttggta
7981 cagcgcagta accggcaaaa tcgcgccgaa ggatgtcgct gcctactggg caatggagcg
8041 cctgccggcc cagtatcagc ccgtcatact tgaagctaga caggcttatc ttggacaaga
8101 agaagatcgc ttggcctcgc gcgcagatca gttggaagaa tttgtccact acgtgaaagg
8161 cgagatcacc aaggtagtcg gcaaataatg tctaacaatt cgttcaagcc gacgccgctt
8221 cgcggcgcgg cttaactcaa gcgttagatg cactaagcac ataattgctc acagccaaac
8281 tatcaggtca agtctgcttt tattattttt aagcgtgcat aataagccct acacaaattg
8341 ggagatatat catgaaaggc tggctttttc ttgttatcgc aatagttggc gaagtaatcg
8401 caacatccgc attaaaatct agcgagggct ttactaagct cctgcagaga tctgaattcc
8461 ctagagagac gaaagtgatt gcgcctaccc ggatattatc gtgaggatgc gtcatcgcca
8521 ttgctcccca aatacaaaac caatttcagc cagtgcctcg tccatttttt cgatgaactc
8581 cggcacgatc tcgtcaaaac tcgccatgta cttttcatcc cgctcaatca cgacataatg
8641 caggccttca cgcttcatac gcgggtcata gttggcaaag taccaggcat tttttcgcgt
8701 cacccacatg ctgtactgca cctgggccat gtaagctgac tttatggcct cgaaaccacc
8761 gagccggaac ttcatgaaat cccgggaggt aaacgggcat ttcagttcaa ggccgttgcc
8821 gtcactgcat aaaccatcgg gagagcaggc ggtacgcata ctttcgtcgc gatagatgat
8881 cggggattca gtaacattca cgccggaagt gaattcaaac agggttctgg cgtcgttctc
8941 gtactgtttt ccccaggcca gtgctttagc gttaacttcc ggagccacac cggtgcaaac
9001 ctcagcaagc agggtgtgga agtaggacat tttcatgtca ggccacttct ttccggagcg
9061 gggttttgct atcacgttgt gaacttctga agcggtgatg acgccgagcc gtaatttgtg
9121 ccacgcatct tccccctgtt cgacagctct cacatcgatc ccggtacgct gcaggataat
9181 gtccggtgtc atgctgccac cttctgctct gcggctttct gtttcaggaa tccaagagct
9241 tttactgctt cggcctgtgt cagttctgac gatgcacgaa tgtcgcggcg aaatatctgg
9301 gaacagagcg gcaataagtc gtcatcccat gttttatcca gggcgatcag cagagtgtta
9361 atctcctgca tggtttcatc gttaaccgga gtgatgtcgc gttccggctg acgttctgca
9421 gtgtatgcag tattttcgac aatgcgctcg gcttcatcct tgtcatagat accagcaaat
9481 ccgaaggcca gacgggcaca ctgaatcatg gctttatgac gtaacatccg tttgggatgc
9541 gactgccacg gccccgtgat ttctctgcct tcgcgagttt tgaatggttc gcggcggcat
9601 tcatccatcc attcggtaac gcagatcgga tgattacggt ccttgcggta aatccggcat
9661 gtacaggatt cattgtcctg ctcaaagtcc atgccatcaa actgctggtt ttcattgatg
9721 atgcgggacc agccatcaac gcccaccacc ggaacgatgc cattctgctt atcaggaaag
9781 gcgtaaattt ctttcgtcca cggattaagg ccgtactggt tggcaacgat cagtaatgcg
9841 atgaactgcg catcgctggc atcaccttta aatgccgtct ggcgaagagt ggtgatcagt
9901 tcctgtgggt cgacagaatc catgccgaca cgttcagcca gcttcccagc cagcgttgcg
9961 agtgcagtac tcattcgttt tatacctctg aatcaatatc aacctggtgg tgagcaatgg
10021 tttcaaccat gtaccggatg tgttctgcca tgcgctcctg aaactcaaca tcgtcatcaa
10081 acgcacgggt aatggatttt ttgctggccc cgtggcgttg caaatgatcg atgcatagcg
10141 attcaaacag gtgctggggc aggccttttt ccatgtcgtc tgccagttct gcctctttct
10201 cttcacgggc gagctgctgg tagtgacgcg cccagctctg agcctcaaga cgatcctgaa
10261 tgtaataagc gttcatggct gaactcctga aatagctgtg aaaatatcgc ccgcgaaatg
10321 ccgggctgat taggaaaaca ggaaaggggg ttagtgaatg cttttgcttg atctcagttt
10381 cagtattaat atccattttt tataagcgtc gactgtttcc tgtgtgaaat tgttatccgc
10441 tcacaattcc acacattata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
10501 gagtgagaat tcggatctcg acgggatgtt gattctgtca tggcatatcc ttacaactta
10561 aaaaagcaaa agggccgcag atgcgaccct tgtgtatcaa acaagacgat taaaaatctt
10621 cgttagtttc tgctacgcct tcgctatcat ctacagagaa atccggcgtt gagttcgggt
10681 tgctcagcag caactcacgt actttcttct cgatctcttt cgcggtttcc gggttatctt
10741 tcagccaggc agtcgcattc gctttaccct gaccgatctt ctcacctttg tagctgtacc
10801 acgcgcctgc tttctcgatc agcttctctt ttacgcccag gtcaaccagt tcgccgtaga
10861 agttgatacc ttcgccgtag aggatctgga attcagcctg tttaaacggc gcagcgattt
10921 tgttcttcac cactttcacg cgggtttcgc tacccaccac gttttcgccc tctttcaccg
10981 cgccgatacg acggatgtcg agacgaacag aggcgtagaa tttcagcgcg ttaccaccgg
11041 tagtggtttc cgggttaccg aacatcacac caattttcat acggatctgg ttgatgaaga
11101 tcagcagcgt gttggactgc ttcaggttac ccgccagctt acgcatcgcc tggctcatca
11161 tacgtgccgc aaggcccatg tgagagtcgc cgatttcgcc ttcgatttcc gctttcggcg
11221 tcagtgccgc cacggagtca acgacgataa cgtctactgc gccagaacgc gccagggcgt
11281 cacagatttc cagtgcctgc tcgccggtgt ccggctggga gcacagcagg ttgtcgatat
11341 cgacgcccag tttacgtgcg tagattgggt ccagcgcgtg ttcagcatcg ataaacgcac
11401 aggttttacc ttcacgctgc gctgcggcga tcacctgcag cgtcagcgtg gttttaccgg
11461 aagattccgg tccgtagatt tcgacgatac ggcccatcgg cagaccacct gccccaagcg
11521 cgatatccag tgaaagcgaa ccggtagaga tggtttccac atccatggaa cggtcttcac
11581 ccaggcgcat gatggagcct ttaccaaatt gtttctcaat ctggcccagt gctgccgcca
11641 acgctttctg tttgttttcg tcgatagcca tttttactcc tgtcatgccg ggtaataccg
11701 gatagtcaat atgttctgtt gaagcaatta tactgtatgc tcatacagta tcaagtgttt
11761 tgtagaaatt gttgccacaa ggtctgcaat gcatacgcag tagcctgacg acgcaccgca
11821 tcacggtcgc cgctgaagca tcccgccggg taatgccttc accgcgggca gtggcaaaag
11881 caaaccagac ggtgccgaca ggcttctcta gatccgccta cctttcacga gttgcgcagt
11941 ttgtctgcaa gactctatga gaagcagata agcgataagt ttgctcaaca tcttctcggg
12001 cataagtcgg acaccatggc atcacagtat cgtgatgaca gaggcaggga gtgggacaaa
12061 attgaaatca aataatgatt ttattttgac tgatagtgac ctgttcgttg caacaaattg
12121 ataagcaatg ccaagcttgg cactggctga tcagctagct cactgcccgc tttccagtcg
12181 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
12241 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
12301 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
12361 gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc
12421 gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg
12481 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
12541 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
12601 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
12661 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
12721 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
12781 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
12841 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
12901 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
12961 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
13021 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
13081 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
13141 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
13201 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
13261 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcaccatt cgatgctagc
13321 ccatgggtat ggacagtttt ccctttgata tgtaacggtg aacagttgtt ctacttttgt
13381 ttgttagtct tgatgcttca ctgatagata caagagccat aagaacctca gatccttccg
13441 tatttagcca gtatgttctc tagtgtggtt cgttgttttt gcgtgagcca tgagaacgaa
13501 ccattgagat catacttact ttgcatgtca ctcaaaaatt ttgcctcaaa actggtgagc
13561 tgaatttttg cagttaaagc atcgtgtagt gtttttctta gtccgttacg taggtaggaa
13621 tctgatgtaa tggttgttgg tattttgtca ccattcattt ttatctggtt gttctcaagt
13681 tcggttacga gatccatttg tctatctagt tcaacttgga aaatcaacgt atcagtcggg
13741 cggcctcgct tatcaaccac caatttcata ttgctgtaag tgtttaaatc tttacttatt
13801 ggtttcaaaa cccattggtt aagcctttta aactcatggt agttattttc aagcattaac
13861 atgaacttaa attcatcaag gctaatctct atatttgcct tgtgagtttt cttttgtgtt
13921 agttctttta ataaccactc ataaatcctc atagagtatt tgttttcaaa agacttaaca
13981 tgttccagat tatattttat gaattttttt aactggaaaa gataaggcaa tatctcttca
14041 ctaaaaacta attctaattt ttcgcttgag aacttggcat agtttgtcca ctggaaaatc
14101 tcaaagcctt taaccaaagg attcctgatt tccacagttc tcgtcatcag ctctctggtt
14161 gctttagcta atacaccata agcattttcc ctactgatgt tcatcatctg agcgtattgg
14221 ttataagtga acgataccgt ccgttctttc cttgtagggt tttcaatcgt ggggttgagt
14281 agtgccacac agcataaaat tagcttggtt tcatgctccg ttaagtcata gcgactaatc
14341 gctagttcat ttgctttgaa aacaactaat tcagacatac atctcaattg gtctaggtga
14401 ttttaatcac tataccaatt gagatgggct agtcaatgat aattactagt ccttttcctt
14461 tgagttgtgg gtatctgtaa attctgctag acctttgctg gaaaacttgt aaattctgct
14521 agaccctctg taaattccgc tagacctttg tgtgtttttt ttgtttatat tcaagtggtt
14581 ataatttata gaataaagaa agaataaaaa aagataaaaa gaatagatcc cagccctgtg
14641 tataactcac tactttagtc agttccgcag tattacaaaa ggatgtcgca aacgctgttt
14701 gctcctctac aaaacagacc ttaaaaccct aaaggcttaa gtagcaccct cgcaagctcg
14761 gttgcggccg caatcgggca aatcgctgaa tattcctttt gtctccgacc atcaggcacc
14821 tgagtcgctg tctttttcgt gacattcagt tcgctgcgct cacggctctg gcagtgaatg
14881 ggggtaaatg gcactacagg cgccttttat ggattcatgc aaggaaacta cccataatac
14941 aagaaaagcc cgtcacgggc ttctcagggc gttttatggc gggtctgcta tgtggtgcta
15001 tctgactttt tgctgttcag cagttcctgc cctctgattt tccagtctga ccacttcgga
15061 ttatcccgtg acaggtcatt cagactggct aatgcaccca gtaaggcagc ggtatcatca
15121 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
15181 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
15241 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
15301 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
15361 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
15421 tcaccggctc cagatttatc agcaataaac cagccagccg
//
