| 质粒名称: | 138-dCas9-Dnmt3a |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 84570 |
| 存储实验室: | Rudolf Jaenisch |
|
载体骨架基本信息 |
|
| 载体名称: | PiggyBac |
| 空载体大小: | 7783 bp |
| 完整质粒大小: | 14723 bp |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 载体抗性: | 氨苄霉素100 μg/mL |
| 生长温度: | 30 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | dCas9-Dnmt3a |
| 物种来源: | H. sapiens (human), Synthetic |
| 插入大小: | 6940 bp |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | NheI(未破坏) |
| 3’克隆位点: | NotI (未破坏) |
| 5’测序引物 | atcgcctggagcaattcc |
|
文献引用方法 |
|
| 材料和方法部分: | 138-dCas9-Dnmt3a was a gift from Rudolf Jaenisch (Addgene plasmid # 84570 ; http://n2t.net/addgene:84570 ; RRID:Addgene_84570) |
| 参考文献部分: | Editing DNA Methylation in the Mammalian Genome. Liu XS, Wu H, Ji X, Stelzer Y, Wu X, Czauderna S, Shu J, Dadon D, Young RA, Jaenisch R. Cell. 2016 Sep 22;167(1):233-247.e17. doi: 10.1016/j.cell.2016.08.056. 10.1016/j.cell.2016.08.056 PubMed 27662091 |
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 138-dCas9-Dnmt3a 14783 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION PiggyBac transposon system construct with dCas9-Dnmt3a.
ACCESSION .
VERSION .
KEYWORDS 138-dCas9-Dnmt3a
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 14783)
AUTHORS Liu XS, Wu H, Ji X, Stelzer Y, Wu X, Czauderna S, Shu J, Dadon D,
Young RA, Jaenisch R
TITLE Editing DNA Methylation in the Mammalian Genome.
JOURNAL Cell. 2016 Sep 22;167(1):233-247.e17. doi:
10.1016/j.cell.2016.08.056.
PUBMED 27662091
REFERENCE 2 (bases 1 to 14783)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..14783
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 46..68
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 60..77
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 61..77
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
repeat_region 207..269
/label=piggyBac right (3') inverted repeat
/note="piggyBac transposon-specific inverted terminal
repeat sequence (ITR)"
promoter 718..1096
/label=TRE3G promoter
/note="3rd-generation Tet-responsive promoter that can be
activated by binding of Tet-On(R) 3G"
protein_bind 726..744
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 762..780
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 798..816
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 834..852
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 870..888
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 906..924
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 942..960
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
primer_bind 1005..1029
/label=LNCX
/note="Human CMV promoter, forward primer"
CDS 1112..5215
/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 5231..5251
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5255..5275
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5279..5299
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5306..8044
/codon_start=1
/product="DNA methyltransferase 3A"
/label=DNMT3A
/translation="MPAMPSSGPGDTSSSAAEREEDRKDGEEQEEPRGKEERQEPSTTA
RKVGRPGRKRKHPPVESGDTPKDPAVISKSPSMAQDSGASELLPNGDLEKRSEPQPEEG
SPAGGQKGGAPAEGEGAAETLPEASRAVENGCCTPKEGRGAPAEAGKEQKETNIESMKM
EGSRGRLRGGLGWESSLRQRPMPRLTFQAGDPYYISKRKRDEWLARWKREAEKKAKVIA
GMNAVEENQGPGESQKVEEASPPAVQQPTDPASPTVATTPEPVGSDAGDKNATKAGDDE
PEYEDGRGFGIGELVWGKLRGFSWWPGRIVSWWMTGRSRAAEGTRWVMWFGDGKFSVVC
VEKLMPLSSFCSAFHQATYNKQPMYRKAIYEVLQVASSRAGKLFPVCHDSDESDTAKAV
EVQNKPMIEWALGGFQPSGPKGLEPPEEEKNPYKEVYTDMWVEPEAAAYAPPPPAKKPR
KSTAEKPKVKEIIDERTRERLVYEVRQKCRNIEDICISCGSLNVTLEHPLFVGGMCQNC
KNCFLECAYQYDDDGYQSYCTICCGGREVLMCGNNNCCRCFCVECVDLLVGPGAAQAAI
KEDPWNCYMCGHKGTYGLLRRREDWPSRLQMFFANNHDQEFDPPKVYPPVPAEKRKPIR
VLSLFDGIATGLLVLKDLGIQVDRYIASEVCEDSITVGMVRHQGKIMYVGDVRSVTQKH
IQEWGPFDLVIGGSPCNDLSIVNPARKGLYEGTGRLFFEFYRLLHDARPKEGDDRPFFW
LFENVVAMGVSDKRDISRFLESNPVMIDAKEVSAAHRARYFWGNLPGMNRPLASTVNDK
LELQECLEHGRIAKFSKVRTITTRSNSIKQGKDQHFPVFMNEKEDILWCTEMERVFGFP
VHYTDVSNMSRLARQRLLGRSWSVPVIRHLFAPLKEYFACV"
primer_bind complement(8095..8114)
/label=TK-pA-R
/note="Thymidine kinase polyA, reverse primer"
polyA_signal 8139..8187
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
promoter 8418..8629
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
LTR 8642..8910
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from human
T-cell leukemia virus (HTLV) type 1"
CDS 8944..9687
/codon_start=1
/product="modified rtTA protein that binds tightly to
promoters containing the tet operator in the presence of
doxycycline"
/label=Tet-On(R) 3G
/translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
DALDDFDLDMLPADALDDFDLDMLPG"
CDS 9688..9741
/codon_start=1
/product="2A peptide from Thosea asigna virus capsid
protein"
/label=T2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="EGRGSLLTCGDVEENPGP"
primer_bind complement(9742..9761)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
CDS 9790..10593
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(9853..9872)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 10463..10482
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 10686..10807
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(10723..10742)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 10777..10796
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
repeat_region 11570..11604
/label=piggyBac left (5') inverted repeat
/note="piggyBac transposon-specific inverted terminal
repeat sequence (ITR)"
primer_bind complement(12095..12111)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(12095..12111)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(12108..12130)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 12119..12135
/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(12143..12173)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 12188..12209
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(12326..12343)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(12497..13085)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(12577..12596)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(13256..14116)
/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 13879..13898
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(14117..14221)
/gene="bla"
/label=AmpR promoter
rep_origin complement(14247..14702)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(14384..14405)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 14596..14615
/label=F1ori-R
/note="F1 origin, reverse primer"
ORIGIN
1 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
61 gtaaaacgac ggccagtgag cgcgcctcgt tcattcacgt ttttgaaccc gtggaggacg
121 ggcagactcg cggtgcaaat gtgttttaca gcgtgatgga gcagatgaag atgctcgaca
181 cgctgcagaa cacgcagcta gattaaccct agaaagataa tcatattgtg acgtacgtta
241 aagataatca tgtgtaaaat tgacgcatgt gttttatcgg tctgtatatc gaggtttatt
301 tattaatttg aatagatatt aagttttatt atatttacac ttacatacta ataataaatt
361 caacaaacaa tttatttatg tttatttatt tattaaaaaa aacaaaaact caaaatttct
421 tctataaagt aacaaaactt ttatgaggga cagccccccc ccaaagcccc cagggatgta
481 attacgtccc tcccccgcta gggggcagca gcgagccgcc cggggctccg ctccggtccg
541 gcgctccccc cgcatccccg agccggcagc gtgcggggac agcccgggca cggggaaggt
601 ggcacgggat cgctttcctc tgaacgcttc tcgctgctct ttgagcctgc agacacctgg
661 ggggatacgg ggaaaaggcc tccacggcca ctagtctttc gtcttcaaga attcctcgag
721 tttactccct atcagtgata gagaacgtat gaagagttta ctccctatca gtgatagaga
781 acgtatgcag actttactcc ctatcagtga tagagaacgt ataaggagtt tactccctat
841 cagtgataga gaacgtatga ccagtttact ccctatcagt gatagagaac gtatctacag
901 tttactccct atcagtgata gagaacgtat atccagttta ctccctatca gtgatagaga
961 acgtataagc tttaggcgtg tacggtgggc gcctataaaa gcagagctcg tttagtgaac
1021 cgtcagatcg cctggagcaa ttccacaaca cttttgtctt ataccaactt tccgtaccac
1081 ttcctaccct cgtaaaggtc tagagctagc catggacaag aagtattcta tcggactggc
1141 catcgggact aatagcgtcg ggtgggccgt gatcactgac gagtacaagg tgccctctaa
1201 gaagttcaag gtgctcggga acaccgaccg gcattccatc aagaaaaatc tgatcggagc
1261 tctcctcttt gattcagggg agaccgctga agcaacccgc ctcaagcgga ctgctagacg
1321 gcggtacacc aggaggaaga accggatttg ttaccttcaa gagatattct ccaacgaaat
1381 ggcaaaggtc gacgacagct tcttccatag gctggaagaa tcattcctcg tggaagagga
1441 taagaagcat gaacggcatc ccatcttcgg taatatcgtc gacgaggtgg cctatcacga
1501 gaaataccca accatctacc atcttcgcaa aaagctggtg gactcaaccg acaaggcaga
1561 cctccggctt atctacctgg ccctggccca catgatcaag ttcagaggcc acttcctgat
1621 cgagggcgac ctcaatcctg acaatagcga tgtggataaa ctgttcatcc agctggtgca
1681 gacttacaac cagctctttg aagagaaccc catcaatgca agcggagtcg atgccaaggc
1741 cattctgtca gcccggctgt caaagagccg cagacttgag aatcttatcg ctcagctgcc
1801 gggtgaaaag aaaaatggac tgttcgggaa cctgattgct ctttcacttg ggctgactcc
1861 caatttcaag tctaatttcg acctggcaga ggatgccaag ctgcaactgt ccaaggacac
1921 ctatgatgac gatctcgaca acctcctggc ccagatcggt gaccaatacg ccgacctttt
1981 ccttgctgct aagaatcttt ctgacgccat cctgctgtct gacattctcc gcgtgaacac
2041 tgaaatcacc aaggcccctc tttcagcttc aatgattaag cggtatgatg agcaccacca
2101 ggacctgacc ctgcttaagg cactcgtccg gcagcagctt ccggagaagt acaaggaaat
2161 cttctttgac cagtcaaaga atggatacgc cggctacatc gacggaggtg cctcccaaga
2221 ggaattttat aagtttatca aacctatcct tgagaagatg gacggcaccg aagagctcct
2281 cgtgaaactg aatcgggagg atctgctgcg gaagcagcgc actttcgaca atgggagcat
2341 tccccaccag atccatcttg gggagcttca cgccatcctt cggcgccaag aggacttcta
2401 cccctttctt aaggacaaca gggagaagat tgagaaaatt ctcactttcc gcatccccta
2461 ctacgtggga cccctcgcca gaggaaatag ccggtttgct tggatgacca gaaagtcaga
2521 agaaactatc actccctgga acttcgaaga ggtggtggac aagggagcca gcgctcagtc
2581 attcatcgaa cggatgacta acttcgataa gaacctcccc aatgagaagg tcctgccgaa
2641 acattccctg ctctacgagt actttaccgt gtacaacgag ctgaccaagg tgaaatatgt
2701 caccgaaggg atgaggaagc ccgcattcct gtcaggcgaa caaaagaagg caattgtgga
2761 ccttctgttc aagaccaata gaaaggtgac cgtgaagcag ctgaaggagg actatttcaa
2821 gaaaattgaa tgcttcgact ctgtggagat tagcggggtc gaagatcggt tcaacgcaag
2881 cctgggtacc taccatgatc tgcttaagat catcaaggac aaggattttc tggacaatga
2941 ggagaacgag gacatccttg aggacattgt cctgactctc actctgttcg aggaccggga
3001 aatgatcgag gagaggctta agacctacgc ccatctgttc gacgataaag tgatgaagca
3061 acttaaacgg agaagatata ccggatgggg acgccttagc cgcaaactca tcaacggaat
3121 ccgggacaaa cagagcggaa agaccattct tgatttcctt aagagcgacg gattcgctaa
3181 tcgcaacttc atgcaactta tccatgatga ttccctgacc tttaaggagg acatccagaa
3241 ggcccaagtg tctggacaag gtgactcact gcacgagcat atcgcaaatc tggctggttc
3301 acccgctatt aagaagggta ttctccagac cgtgaaagtc gtggacgagc tggtcaaggt
3361 gatgggtcgc cataaaccag agaacattgt catcgagatg gccagggaaa accagactac
3421 ccagaaggga cagaagaaca gcagggagcg gatgaaaaga attgaggaag ggattaagga
3481 gctcgggtca cagatcctta aagagcaccc ggtggaaaac acccagcttc agaatgagaa
3541 gctctatctg tactaccttc aaaatggacg cgatatgtat gtggaccaag agcttgatat
3601 caacaggctc tcagactacg acgtggacgc catcgtccct cagagcttcc tcaaagacga
3661 ctcaattgac aataaggtgc tgactcgctc agacaagaac cggggaaagt cagataacgt
3721 gccctcagag gaagtcgtga aaaagatgaa gaactattgg cgccagcttc tgaacgcaaa
3781 gctgatcact cagcggaagt tcgacaatct cactaaggct gagaggggcg gactgagcga
3841 actggacaaa gcaggattca ttaaacggca acttgtggag actcggcaga ttactaaaca
3901 tgtcgcccaa atccttgact cacgcatgaa taccaagtac gacgaaaacg acaaacttat
3961 ccgcgaggtg aaggtgatta ccctgaagtc caagctggtc agcgatttca gaaaggactt
4021 tcaattctac aaagtgcggg agatcaataa ctatcatcat gctcatgacg catatctgaa
4081 tgccgtggtg ggaaccgccc tgatcaagaa gtacccaaag ctggaaagcg agttcgtgta
4141 cggagactac aaggtctacg acgtgcgcaa gatgattgcc aaatctgagc aggagatcgg
4201 aaaggccacc gcaaagtact tcttctacag caacatcatg aatttcttca agaccgaaat
4261 cacccttgca aacggtgaga tccggaagag gccgctcatc gagactaatg gggagactgg
4321 cgaaatcgtg tgggacaagg gcagagattt cgctaccgtg cgcaaagtgc tttctatgcc
4381 tcaagtgaac atcgtgaaga aaaccgaggt gcaaaccgga ggcttttcta aggaatcaat
4441 cctccccaag cgcaactccg acaagctcat tgcaaggaag aaggattggg accctaagaa
4501 gtacggcgga ttcgattcac caactgtggc ttattctgtc ctggtcgtgg ctaaggtgga
4561 aaaaggaaag tctaagaagc tcaagagcgt gaaggaactg ctgggtatca ccattatgga
4621 gcgcagctcc ttcgagaaga acccaattga ctttctcgaa gccaaaggtt acaaggaagt
4681 caagaaggac cttatcatca agctcccaaa gtatagcctg ttcgaactgg agaatgggcg
4741 gaagcggatg ctcgcctccg ctggcgaact tcagaagggt aatgagctgg ctctcccctc
4801 caagtacgtg aatttcctct accttgcaag ccattacgag aagctgaagg ggagccccga
4861 ggacaacgag caaaagcaac tgtttgtgga gcagcataag cattatctgg acgagatcat
4921 tgagcagatt tccgagtttt ctaaacgcgt cattctcgct gatgccaacc tcgataaagt
4981 ccttagcgca tacaataagc acagagacaa accaattcgg gagcaggctg agaatatcat
5041 ccacctgttc accctcacca atcttggtgc ccctgccgca ttcaagtact tcgacaccac
5101 catcgaccgg aaacgctata cctccaccaa agaagtgctg gacgccaccc tcatccacca
5161 gagcatcacc ggactttacg aaactcggat tgacctctca cagctcggag gggatggtgg
5221 cggaggctcg ccaaaaaaga agagaaaggt agacccaaag aaaaaacgaa aagtagatcc
5281 gaaaaagaag aggaaggtgg gatccatgcc cgccatgccc tccagcggcc ccggggacac
5341 cagcagctct gctgcggagc gggaggagga ccgaaaggac ggagaggagc aggaggagcc
5401 gcgtggcaag gaggagcgcc aagagcccag caccacggca cggaaggtgg ggcggcctgg
5461 gaggaagcgc aagcaccccc cggtggaaag cggtgacacg ccaaaggacc ctgcggtgat
5521 ctccaagtcc ccatccatgg cccaggactc aggcgcctca gagctattac ccaatgggga
5581 cttggagaag cggagtgagc cccagccaga ggaggggagc cctgctgggg ggcagaaggg
5641 cggggcccca gcagagggag agggtgcagc tgagaccctg cctgaagcct caagagcagt
5701 ggaaaatggc tgctgcaccc ccaaggaggg ccgaggagcc cctgcagaag cgggcaaaga
5761 acagaaggag accaacatcg aatccatgaa aatggagggc tcccggggcc ggctgcgggg
5821 tggcttgggc tgggagtcca gcctccgtca gcggcccatg ccgaggctca ccttccaggc
5881 gggggacccc tactacatca gcaagcgcaa gcgggacgag tggctggcac gctggaaaag
5941 ggaggctgag aagaaagcca aggtcattgc aggaatgaat gctgtggaag aaaaccaggg
6001 gcccggggag tctcagaagg tggaggaggc cagccctcct gctgtgcagc agcccactga
6061 ccccgcatcc cccactgtgg ctaccacgcc tgagcccgtg gggtccgatg ctggggacaa
6121 gaatgccacc aaagcaggcg atgacgagcc agagtacgag gacggccggg gctttggcat
6181 tggggagctg gtgtggggga aactgcgggg cttctcctgg tggccaggcc gcattgtgtc
6241 ttggtggatg acgggccgga gccgagcagc tgaaggcacc cgctgggtca tgtggttcgg
6301 agacggcaaa ttctcagtgg tgtgtgttga gaagctgatg ccgctgagct cgttttgcag
6361 tgcgttccac caggccacgt acaacaagca gcccatgtac cgcaaagcca tctacgaggt
6421 cctgcaggtg gccagcagcc gcgcggggaa gctgttcccg gtgtgccacg acagcgatga
6481 gagtgacact gccaaggccg tggaggtgca gaacaagccc atgattgaat gggccctggg
6541 gggcttccag ccttctggcc ctaagggcct ggagccacca gaagaagaga agaatcccta
6601 caaagaagtg tacacggaca tgtgggtgga acctgaggca gctgcctacg caccacctcc
6661 accagccaaa aagccccgga agagcacagc ggagaagccc aaggtcaagg agattattga
6721 tgagcgcaca agagagcggc tggtgtacga ggtgcggcag aagtgccgga acattgagga
6781 catctgcatc tcctgtggga gcctcaatgt taccctggaa caccccctct tcgttggagg
6841 aatgtgccaa aactgcaaga actgctttct ggagtgtgcg taccagtacg acgacgacgg
6901 ctaccagtcc tactgcacca tctgctgtgg gggccgtgag gtgctcatgt gcggaaacaa
6961 caactgctgc aggtgctttt gcgtggagtg tgtggacctc ttggtggggc cgggggctgc
7021 ccaggcagcc attaaggaag acccctggaa ctgctacatg tgcgggcaca agggtaccta
7081 cgggctgctg cggcggcgag aggactggcc ctcccggctc cagatgttct tcgctaataa
7141 ccacgaccag gaatttgacc ctccaaaggt ttacccacct gtcccagctg agaagaggaa
7201 gcccatccgg gtgctgtctc tctttgatgg aatcgctaca gggctcctgg tgctgaagga
7261 cttgggcatt caggtggacc gctacattgc ctcggaggtg tgtgaggact ccatcacggt
7321 gggcatggtg cggcaccagg ggaagatcat gtacgtcggg gacgtccgca gcgtcacaca
7381 gaagcatatc caggagtggg gcccattcga tctggtgatt gggggcagtc cctgcaatga
7441 cctctccatc gtcaaccctg ctcgcaaggg cctctacgag ggcactggcc ggctcttctt
7501 tgagttctac cgcctcctgc atgatgcgcg gcccaaggag ggagatgatc gccccttctt
7561 ctggctcttt gagaatgtgg tggccatggg cgttagtgac aagagggaca tctcgcgatt
7621 tctcgagtcc aaccctgtga tgattgatgc caaagaagtg tcagctgcac acagggcccg
7681 ctacttctgg ggtaaccttc ccggtatgaa caggccgttg gcatccactg tgaatgataa
7741 gctggagctg caggagtgtc tggagcatgg caggatagcc aagttcagca aagtgaggac
7801 cattactacg aggtcaaact ccataaagca gggcaaagac cagcattttc ctgtcttcat
7861 gaatgagaaa gaggacatct tatggtgcac tgaaatggaa agggtatttg gtttcccagt
7921 ccactatact gacgtctcca acatgagccg cttggcgagg cagagactgc tgggccggtc
7981 atggagcgtg ccagtcatcc gccacctctt cgctccgctg aaggagtatt ttgcgtgtgt
8041 gtaagcggcc gcatatatac cggttagtaa tgagtttaaa cgggggaggc taactgaaac
8101 acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa
8161 aacgcacggg tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact
8221 ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc
8281 caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag
8341 gccctgccat agcagatctg cgaaatcgga tccagctgaa tctaagtgga tctgcgatcg
8401 ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg
8461 aggggtcggc aattgaacgg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga
8521 tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt
8581 agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagc tgaagcttcg
8641 aggggctcgc atctctcctt cacgcgcccg ccgccctacc tgaggccgcc atccacgccg
8701 gttgagtcgc gttctgccgc ctcccgcctg tggtgcctcc tgaactgcgt ccgccgtcta
8761 ggtaagttta aagctcaggt cgagaccggg cctttgtccg gcgctccctt ggagcctacc
8821 tagactcagc cggctctcca cgctttgcct gaccctgctt gctcaactct acgtctttgt
8881 ttcgttttct gttctgcgcc gttacagatc taagctgtga ccggcgccta ctctagagcc
8941 gccatgtcta gactggacaa gagcaaagtc ataaactctg ctctggaatt actcaatgga
9001 gtcggtatcg aaggcctgac gacaaggaaa ctcgctcaaa agctgggagt tgagcagcct
9061 accctgtact ggcacgtgaa gaacaagcgg gccctgctcg atgccctgcc aatcgagatg
9121 ctggacaggc atcataccca ctcctgcccc ctggaaggcg agtcatggca agactttctg
9181 cggaacaacg ccaagtcata ccgctgtgct ctcctctcac atcgcgacgg ggctaaagtg
9241 catctcggca cccgcccaac agagaaacag tacgaaaccc tggaaaatca gctcgcgttc
9301 ctgtgtcagc aaggcttctc cctggagaac gcactgtacg ctctgtccgc cgtgggccac
9361 tttacactgg gctgcgtatt ggaggaacag gagcatcaag tagcaaaaga ggaaagagag
9421 acacctacca ccgattctat gcccccactt ctgaaacaag caattgagct gttcgaccgg
9481 cagggagccg aacctgcctt ccttttcggc ctggaactaa tcatatgtgg cctggagaaa
9541 cagctaaagt gcgaaagcgg cgggccgacc gacgcccttg acgattttga cttagacatg
9601 ctcccagccg atgcccttga cgactttgac cttgatatgc tgcctgctga cgctcttgac
9661 gattttgacc ttgacatgct ccccggggag ggcagaggaa gtcttctaac atgcggtgac
9721 gtggaggaga atcccggccc tatgaccgag tacaagccca cggtgcgcct cgccacccgc
9781 gacgacgtca tgggatcggc cattgaacaa gatggattgc acgcaggttc tccggccgct
9841 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
9901 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
9961 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
10021 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
10081 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
10141 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
10201 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
10261 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
10321 aaggcgcgca tgcccgacgg cgatgatctc gtcgtgaccc atggcgatgc ctgcttgccg
10381 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
10441 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
10501 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
10561 gccttctatc gccttcttga cgagttcttc tgagacgtcg aggtgcccga aggaccgcgc
10621 acctggtgca tgacccgcaa gcccggtgcc tgaaatcaac ctctggatta caaaatttgt
10681 gggtaaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
10741 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
10801 tatcttatca tgtctggaat tgactcaaat gatgtcaatt agtctatcag aagctcatct
10861 ggtctccctt ccgggggaca agacatccct gtttaatatt taaacagcag tgttcccaaa
10921 ctgggttctt atatcccttg ctctggtcaa ccaggttgca gggtttcctg tcctcacagg
10981 aacgaagtcc ctaaagaaac agtggcagcc aggtttagcc ccggaattga ctggattcct
11041 tttttagggc ccattggtat ggctttttcc ccgtatcccc ccaggtgtct gcaggctcaa
11101 agagcagcga gaagcgttca gaggaaagcg atcccgtgcc accttccccg tgcccgggct
11161 gtccccgcac gctgccggct cggggatgcg gggggagcgc cggaccggag cggagccccg
11221 ggcggctcgc tgctgccccc tagcggggga gggacgtaat tacatccctg ggggctttgg
11281 gggggggctg tccctgatat ctataacaag aaaatatata tataataagt tatcacgtaa
11341 gtagaacatg aaataacaat ataattatcg tatgagttaa atcttaaaag tcacgtaaaa
11401 gataatcatg cgtcattttg actcacgcgg tcgttatagt tcaaaatcag tgacacttac
11461 cgcattgaca agcacgcctc acgggagctc caagcggcga ctgagatgtc ctaaatgcac
11521 agcgacggat tcgcgctatt tagaaagaga gagcaatatt tcaagaatgc atgcgtcaat
11581 tttacgcaga ctatctttct agggttaatc tagctgcatc aggatcatat cgtcgggtct
11641 tttttccggc tcagtcatcg cccaagctgg cgctatctgg gcatcgggga ggaagaagcc
11701 cgtgcctttt cccgcgaggt tgaagcggca tggaaagagt ttgccgagga tgactgctgc
11761 tgcattgacg ttgagcgaaa acgcacgttt accatgatga ttcgggaagg tgtggccatg
11821 cacgccttta acggtgaact gttcgttcag gccacctggg ataccagttc gtcgcggctt
11881 ttccggacac agttccggat ggtcagcccg aagcgcatca gcaacccgaa caataccggc
11941 gacagccgga actgccgtgc cggtgtgcag attaatgaca gcggtgcggc gctgggatat
12001 tacgtcagcg aggacgggta tcctggctgg atgccgcaga aatggacatg gataccccgt
12061 gagttacccg gcgggcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
12121 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
12181 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
12241 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
12301 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
12361 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
12421 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
12481 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
12541 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
12601 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
12661 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
12721 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
12781 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
12841 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
12901 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
12961 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
13021 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
13081 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
13141 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
13201 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
13261 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
13321 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
13381 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
13441 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
13501 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
13561 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
13621 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
13681 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
13741 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
13801 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
13861 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
13921 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
13981 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
14041 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
14101 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
14161 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
14221 gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc
14281 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
14341 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
14401 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
14461 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
14521 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
14581 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
14641 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
14701 gtcccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
14761 gctattacgc cagctggcga aag
//
