| 质粒名称: | pCRCT |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 60621 |
| 存储实验室: | Huimin Zhao |
|
载体骨架基本信息 |
|
| 载体名称: | pRS426 |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 5000 bp |
| 完整质粒大小: | 10805 bp |
| 载体修饰: | URA3启动子被截断 |
| 载体类型: | 酵母表达，CRISPR |
| 筛选标记: | URA3 |
| 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: | iCas9 |
| 物种来源: | 化脓性链球菌 |
| 插入大小: | 4179 bp |
| 基因突变: | 将天冬氨酸147改为酪氨酸，脯氨酸411改为苏氨酸 |
| GenBank编号 | -- |
| 启动子 | TEF1p |
| 标签/融合蛋白: |
标志（插入时N端子）
SV40 NLS（N端子插入）
SV40 NLS（C端子插入）
|
| 克隆方法: | 非连接克隆 |
| 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: | pCRCT was a gift from Huimin Zhao (Addgene plasmid # 60621 ; http://n2t.net/addgene:60621 ; RRID:Addgene_60621) |
| 参考文献部分: | Homology-Integrated CRISPR-Cas (HI-CRISPR) System for One-Step Multigene Disruption in Saccharomyces cerevisiae. Bao Z, Xiao H, Liang J, Zhang L, Xiong X, Sun N, Si T, Zhao H. ACS Synth Biol. 2014 Sep 19. 10.1021/sb500255k PubMed 25207793 |
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 pCRCT 10807 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Plasmid encoding iCas9, tracrRNA and crRNAs.
ACCESSION .
VERSION .
KEYWORDS pCRCT
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 10807)
AUTHORS Bao Z, Xiao H, Liang J, Zhang L, Xiong X, Sun N, Si T, Zhao H
TITLE Homology-Integrated CRISPR-Cas (HI-CRISPR) System for One-Step
Multigene Disruption in Saccharomyces cerevisiae.
JOURNAL ACS Synth Biol. 2014 Sep 19.
PUBMED 25207793
REFERENCE 2 (bases 1 to 10807)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..10807
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 54..857
/codon_start=1
/gene="S. cerevisiae URA3"
/product="orotidine-5'-phosphate decarboxylase, required
for uracil biosynthesis"
/label=URA3
/note="yeast auxotrophic marker, counterselectable with
5-fluoroorotic acid (5-FOA)"
/translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
promoter 946..1214
/label=SNR52 promoter
/note="promoter for the S. cerevisiae small nucleolar RNA
gene SNR52"
CDS complement(1257..1553)
/codon_start=1
/gene="lacZ fragment"
/product="LacZ-alpha fragment of beta-galactosidase"
/label=lacZ-alpha
/translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSG"
primer_bind 1403..1425
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 1417..1434
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 1418..1434
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 1444..1463
/label=T7
/note="T7 promoter, forward primer"
promoter 1444..1462
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(1510..1530)
/label=T3
/note="T3 promoter, forward primer"
promoter complement(1510..1528)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(1549..1565)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 1573..1589
/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(1597..1627)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 1642..1663
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
terminator 1736..1755
/label=SUP4 terminator
/note="transcription terminator for the S. cerevisiae SUP4
tRNA gene"
promoter 1757..2156
/gene="S. cerevisiae TEF1"
/label=TEF1 promoter
/note="promoter for EF-1-alpha"
CDS 2168..2191
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
CDS 2195..2215
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 2216..6316
/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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTYKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSITHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 6320..6340
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
terminator 6344..6643
/label=ADH2 terminator
/note="transcription terminator for the S. cerevisiae
alcohol dehydrogenase 2 (ADH2) gene"
promoter 6656..7062
/gene="RPR1"
/label=RPR1 promoter
/note="promoter for the S. cerevisiae RNase P RNA gene"
misc_RNA 7063..7141
/label=tracrRNA
/note="trans-activating CRISPR RNA for the Streptococcus
pyogenes CRISPR/Cas9 system"
terminator 7149..7211
/label=RPR1 terminator
/note="transcription terminator for the S. cerevisiae RNase
P RNA gene"
primer_bind complement(7244..7260)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 7268..7284
/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(7292..7322)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 7337..7358
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(7475..7492)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(7646..8234)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(7726..7745)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(8405..9265)
/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 9028..9047
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(9266..9370)
/gene="bla"
/label=AmpR promoter
rep_origin 9397..10739
/label=2u ori
/note="yeast 2u plasmid origin of replication"
protein_bind complement(10360..10407)
/label=FRT
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
primer_bind 10780..10798
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
ORIGIN
1 tcgcgcgaac ccaactgcac agaacaaaaa cctgcaggaa acgaagataa atcatgtcga
61 aagctacata taaggaacgt gctgctactc atcctagtcc tgttgctgcc aagctattta
121 atatcatgca cgaaaagcaa acaaacttgt gtgcttcatt ggatgttcgt accaccaagg
181 aattactgga gttagttgaa gcattaggtc ccaaaatttg tttactaaaa acacatgtgg
241 atatcttgac tgatttttcc atggagggca cagttaagcc gctaaaggca ttatccgcca
301 agtacaattt tttactcttc gaagacagaa aatttgctga cattggtaat acagtcaaat
361 tgcagtactc tgcgggtgta tacagaatag cagaatgggc agacattacg aatgcacacg
421 gtgtggtggg cccaggtatt gttagcggtt tgaagcaggc ggcagaagaa gtaacaaagg
481 aacctagagg ccttttgatg ttagcagaat tgtcatgcaa gggctcccta tctactggag
541 aatatactaa gggtactgtt gacattgcga agagcgacaa agattttgtt atcggcttta
601 ttgctcaaag agacatgggt ggaagagatg aaggttacga ttggttgatt atgacacccg
661 gtgtgggttt agatgacaag ggagacgcat tgggtcaaca gtatagaacc gtggatgatg
721 tcgtttctac aggatctgac attattattg ttggaagagg actatttgca aagggaaggg
781 atgctaaggt agagggtgaa cgttacagaa aagcaggctg ggaagcatat ttgagaagat
841 gcggccagca aaactaaaaa actgtattat aagtaaatgc atgtatacta aactcacaaa
901 ttagagcttc aatttaatta tatcagttat taccctgtgc tcgagtcttt gaaaagataa
961 tgtatgatta tgctttcact catatttata cagaaacttg atgttttctt tcgagtatat
1021 acaaggtgat tacatgtacg tttgaagtac aactctagat tttgtagtgc cctcttgggc
1081 tagcggtaaa ggtgcgcatt ttttcacacc ctacaatgtt ctgttcaaaa gattttggtc
1141 aaacgctgta gaagtgaaag ttggtgcgca tgtttcggcg ttcgaaactt ctccgcagtg
1201 aaagataaat gatcgggttt tagagctatg ctgttttgaa tggtcccaaa gagacctcag
1261 ccgctacagg gcgcgtccca ttcgccattc aggctgcgca actgttggga agggcgatcg
1321 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta
1381 agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgagcgc
1441 gcgtaatacg actcactata gggcgaattg ggtaccgggc cccccctcga ggtcctccag
1501 cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt
1561 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
1621 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
1681 gcccgctttc caccggtggt ctctagagct atgctgtttt gaatggtccc aaaacttttt
1741 ttgtttttta tgtctgcata gcttcaaaat gtttctactc cttttttact cttccagatt
1801 ttctcggact ccgcgcatcg ccgtaccact tcaaaacacc caagcacagc atactaaatt
1861 tcccctcttt cttcctctag ggtgtcgtta attacccgta ctaaaggttt ggaaaagaaa
1921 aaagaggccg cctcgtttct ttttcttcgt cgaaaaaggc aataaaaatt tttatcacgt
1981 ttctttttct tgaaaatttt tttttttgat ttttttctct ttcgatgacc tcccattgat
2041 atttaagtta ataaacggtc ttcaatttct caagtttcag tttcattttt cttgttctat
2101 tacaactttt tttacttctt gctcattaga aagaaagcat agcaatctaa tctaagtttt
2161 ctagatggat tataaagatg acgatgacaa acctccaaaa aagaagagaa aggtcgataa
2221 gaaatactca ataggcttag atatcggcac aaatagcgtc ggatgggcgg tgatcactga
2281 tgaatataag gttccgtcta aaaagttcaa ggttctggga aatacagacc gccacagtat
2341 caaaaaaaat cttatagggg ctcttttatt tgacagtgga gagacagcgg aagcgactcg
2401 tctcaaacgg acagctcgta gaaggtatac acgtcggaag aatcgtattt gttatctaca
2461 ggagattttt tcaaatgaga tggcgaaagt agatgatagt ttctttcatc gacttgaaga
2521 gtcttttttg gtggaagaag acaagaagca tgaacgtcat cctatttttg gaaatatagt
2581 agatgaagtt gcttatcatg agaaatatcc aactatctat catctgcgaa aaaaattggt
2641 agattctact tataaagcgg atttgcgctt aatctatttg gccttagcgc atatgattaa
2701 gtttcgtggt cattttttga ttgagggaga tttaaatcct gataatagtg atgtggacaa
2761 actatttatc cagttggtac aaacctacaa tcaattattt gaagaaaacc ctattaacgc
2821 aagtggagta gatgctaaag cgattctttc tgcacgattg agtaaatcaa gacgattaga
2881 aaatctcatt gctcagctcc ccggtgagaa gaaaaatggc ttatttggga atctcattgc
2941 tttgtcattg ggtttgaccc ctaattttaa atcaaatttt gatttggcag aagatgctaa
3001 attacagctt tcaaaagata cttacgatga tgatttagat aatttattgg cgcaaattgg
3061 agatcaatat gctgatttgt ttttggcagc taagaattta tcagatgcta ttttactttc
3121 agatatccta agagtaaata ctgaaataac taaggctccc ctatcagctt caatgattaa
3181 acgctacgat gaacatcatc aagacttgac tcttttaaaa gctttagttc gacaacaact
3241 tccagaaaag tataaagaaa tcttttttga tcaatcaaaa aacggatatg caggttatat
3301 tgatggggga gctagccaag aagaatttta taaatttatc aaaccaattt tagaaaaaat
3361 ggatggtact gaggaattat tggtgaaact aaatcgtgaa gatttgctgc gcaagcaacg
3421 gacctttgac aacggctcta ttacccatca aattcacttg ggtgagctgc atgctatttt
3481 gagaagacaa gaagactttt atccattttt aaaagacaat cgtgagaaga ttgaaaaaat
3541 cttgactttt cgaattcctt attatgttgg tccattggcg cgtggcaata gtcgttttgc
3601 atggatgact cggaagtctg aagaaacaat taccccatgg aattttgaag aagttgtcga
3661 taaaggtgct tcagctcaat catttattga acgcatgaca aactttgata aaaatcttcc
3721 aaatgaaaaa gtactaccaa aacatagttt gctttatgag tattttacgg tttataacga
3781 attgacaaag gtcaaatatg ttactgaagg aatgcgaaaa ccagcatttc tttcaggtga
3841 acagaagaaa gccattgttg atttactctt caaaacaaat cgaaaagtaa ccgttaagca
3901 attaaaagaa gattatttca aaaaaataga atgttttgat agtgttgaaa tttcaggagt
3961 tgaagataga tttaatgctt cattaggtac ctaccatgat ttgctaaaaa ttattaaaga
4021 taaagatttt ttggataatg aagaaaatga agatatctta gaggatattg ttttaacatt
4081 gaccttattt gaagataggg agatgattga ggaaagactt aaaacatatg ctcacctctt
4141 tgatgataag gtgatgaaac agcttaaacg tcgccgttat actggttggg gacgtttgtc
4201 tcgaaaattg attaatggta ttagggataa gcaatctggc aaaacaatat tagatttttt
4261 gaaatcagat ggttttgcca atcgcaattt tatgcagctg atccatgatg atagtttgac
4321 atttaaagaa gacattcaaa aagcacaagt gtctggacaa ggcgatagtt tacatgaaca
4381 tattgcaaat ttagctggta gccctgctat taaaaaaggt attttacaga ctgtaaaagt
4441 tgttgatgaa ttggtcaaag taatggggcg gcataagcca gaaaatatcg ttattgaaat
4501 ggcacgtgaa aatcagacaa ctcaaaaggg ccagaaaaat tcgcgagagc gtatgaaacg
4561 aatcgaagaa ggtatcaaag aattaggaag tcagattctt aaagagcatc ctgttgaaaa
4621 tactcaattg caaaatgaaa agctctatct ctattatctc caaaatggaa gagacatgta
4681 tgtggaccaa gaattagata ttaatcgttt aagtgattat gatgtcgatc acattgttcc
4741 acaaagtttc cttaaagacg attcaataga caataaggtc ttaacgcgtt ctgataaaaa
4801 tcgtggtaaa tcggataacg ttccaagtga agaagtagtc aaaaagatga aaaactattg
4861 gagacaactt ctaaacgcca agttaatcac tcaacgtaag tttgataatt taacgaaagc
4921 tgaacgtgga ggtttgagtg aacttgataa agctggtttt atcaaacgcc aattggttga
4981 aactcgccaa atcactaagc atgtggcaca aattttggat agtcgcatga atactaaata
5041 cgatgaaaat gataaactta ttcgagaggt taaagtgatt accttaaaat ctaaattagt
5101 ttctgacttc cgaaaagatt tccaattcta taaagtacgt gagattaaca attaccatca
5161 tgcccatgat gcgtatctaa atgccgtcgt tggaactgct ttgattaaga aatatccaaa
5221 acttgaatcg gagtttgtct atggtgatta taaagtttat gatgttcgta aaatgattgc
5281 taagtctgag caagaaatag gcaaagcaac cgcaaaatat ttcttttact ctaatatcat
5341 gaacttcttc aaaacagaaa ttacacttgc aaatggagag attcgcaaac gccctctaat
5401 cgaaactaat ggggaaactg gagaaattgt ctgggataaa gggcgagatt ttgccacagt
5461 gcgcaaagta ttgtccatgc cccaagtcaa tattgtcaag aaaacagaag tacagacagg
5521 cggattctcc aaggagtcaa ttttaccaaa aagaaattcg gacaagctta ttgctcgtaa
5581 aaaagactgg gatccaaaaa aatatggtgg ttttgatagt ccaacggtag cttattcagt
5641 cctagtggtt gctaaggtgg aaaaagggaa atcgaagaag ttaaaatccg ttaaagagtt
5701 actagggatc acaattatgg aaagaagttc ctttgaaaaa aatccgattg actttttaga
5761 agctaaagga tataaggaag ttaaaaaaga cttaatcatt aaactaccta aatatagtct
5821 ttttgagtta gaaaacggtc gtaaacggat gctggctagt gccggagaat tacaaaaagg
5881 aaatgagctg gctctgccaa gcaaatatgt gaatttttta tatttagcta gtcattatga
5941 aaagttgaag ggtagtccag aagataacga acaaaaacaa ttgtttgtgg agcagcataa
6001 gcattattta gatgagatta ttgagcaaat cagtgaattt tctaagcgtg ttattttagc
6061 agatgccaat ttagataaag ttcttagtgc atataacaaa catagagaca aaccaatacg
6121 tgaacaagca gaaaatatta ttcatttatt tacgttgacg aatcttggag ctcccgctgc
6181 ttttaaatat tttgatacaa caattgatcg taaacgatat acgtctacaa aagaagtttt
6241 agatgccact cttatccatc aatccatcac tggtctttat gaaacacgca ttgatttgag
6301 tcagctagga ggtgaccctc caaaaaagaa gagaaaggtc tgagcggatc tcttatgtct
6361 ttacgattta tagttttcat tatcaagtat gcctatatta gtatatagca tctttagatg
6421 acagtgttcg aagtttcacg aataaaagat aatattctac tttttgctcc caccgcgttt
6481 gctagcacga gtgaacacca tccctcgcct gtgagttgta cccattcctc taaactgtag
6541 acatggtagc ttcagcagtg ttcgttatgt acggcatcct ccaacaaaca gtcggttata
6601 gtttgtcctg ctcctctgaa tcgagtccct cgatatttct catactagtt ctagagatct
6661 gccaattgaa cataacatgg tagttacata tactagtaat atggttcggc acacattaaa
6721 agtataaaaa ctatctgaat tacgaattac atatattggt cataaaaatc aatcaatcat
6781 cgtgtgtttt atatgtctct tatctaagta taagaatatc catagttaat attcacttac
6841 gctacctttt aacctgtaat cattgtcaac aggatatgtt aacgacccac attgataaac
6901 gctagtattt ctttttcctc ttcttattgg ccggctgtct ctatactccc ctatagtctg
6961 tttcttttcg tttcgattgt tttacgtttg aggcctcgtg gcgcacatgg tacgctgtgg
7021 tgctcgcggc tgggaacgaa actctgggag ctgcgattgg caggaaccat tcaaaacagc
7081 atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg
7141 ctttttttcc atatccaact tccaatttaa tctttctttt ttaattttca cttatttgcg
7201 atacagaaag accctgcagg catgcaagct tggcgtaatc atggtcatag ctgtttcctg
7261 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
7321 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
7381 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
7441 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
7501 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
7561 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
7621 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
7681 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
7741 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
7801 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
7861 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
7921 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
7981 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
8041 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
8101 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
8161 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
8221 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
8281 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
8341 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
8401 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
8461 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
8521 gccccagtgc tgcaatgata ccgcggctcc cacgctcacc ggctccagat ttatcagcaa
8581 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
8641 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
8701 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
8761 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
8821 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
8881 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
8941 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
9001 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
9061 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
9121 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
9181 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
9241 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
9301 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
9361 gggttccgcg cacatttccc cgaaaagtgc cacctgaacg aagcatctgt gcttcatttt
9421 gtagaacaaa aatgcaacgc gagagcgcta atttttcaaa caaagaatct gagctgcatt
9481 tttacagaac agaaatgcaa cgcgaaagcg ctattttacc aacgaagaat ctgtgcttca
9541 tttttgtaaa acaaaaatgc aacgcgagag cgctaatttt tcaaacaaag aatctgagct
9601 gcatttttac agaacagaaa tgcaacgcga gagcgctatt ttaccaacaa agaatctata
9661 cttctttttt gttctacaaa aatgcatccc gagagcgcta tttttctaac aaagcatctt
9721 agattacttt ttttctcctt tgtgcgctct ataatgcagt ctcttgataa ctttttgcac
9781 tgtaggtccg ttaaggttag aagaaggcta ctttggtgtc tattttctct tccataaaaa
9841 aagcctgact ccacttcccg cgtttactga ttactagcga agctgcgggt gcattttttc
9901 aagataaagg catccccgat tatattctat accgatgtgg attgcgcata ctttgtgaac
9961 agaaagtgat agcgttgatg attcttcatt ggtcagaaaa ttatgaacgg tttcttctat
10021 tttgtctcta tatactacgt ataggaaatg tttacatttt cgtattgttt tcgattcact
10081 ctatgaatag ttcttactac aatttttttg tctaaagagt aatactagag ataaacataa
10141 aaaatgtaga ggtcgagttt agatgcaagt tcaaggagcg aaaggtggat gggtaggtta
10201 tatagggata tagcacagag atatatagca aagagatact tttgagcaat gtttgtggaa
10261 gcggtattcg caatatttta gtagctcgtt acagtccggt gcgtttttgg ttttttgaaa
10321 gtgcgtcttc agagcgcttt tggttttcaa aagcgctctg aagttcctat actttctaga
10381 gaataggaac ttcggaatag gaacttcaaa gcgtttccga aaacgagcgc ttccgaaaat
10441 gcaacgcgag ctgcgcacat acagctcact gttcacgtcg cacctatatc tgcgtgttgc
10501 ctgtatatat atatacatga gaagaacggc atagtgcgtg tttatgctta aatgcgtact
10561 tatatgcgtc tatttatgta ggatgaaagg tagtctagta cctcctgtga tattatccca
10621 ttccatgcgg ggtatcgtat gcttccttca gcactaccct ttagctgttc tatatgctgc
10681 cactcctcaa ttggattagt ctcatccttc aatgctatca tttcctttga tattggatca
10741 tactaagaaa ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc
10801 tttcgtc
//
