| 质粒名称: | lentiCas9-EGFP |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 63592 |
| 存储实验室: | Phil Sharp, Feng Zhang |
|
载体骨架基本信息 |
|
| 载体名称: | pFUGW |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | 13178bp |
| 载体修饰: | 人细胞色素c氧化酶亚基VIII（human cytochrome C oxidase subunit VIII） |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 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: | Cas9 |
| 物种来源: | -- |
| 插入大小: | 4200 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | 人细胞色素c氧化酶亚基VIII（N端） |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | AgeI(未破坏) |
| 3' 克隆位点: | NotI (未破坏) |
| 5' 测序引物 | CMV-F |
| 3' 测序引物 | SV40pA-R |
|
文献引用方法 |
|
| 材料和方法部分: | lentiCas9-EGFP was a gift from Phil Sharp & Feng Zhang (Addgene plasmid # 63592 ; http://n2t.net/addgene:63592 ; RRID:Addgene_63592) |
| 参考文献部分: | Genome-wide CRISPR screen in a mouse model of tumor growth and metastasis. Chen S, Sanjana NE, Zheng K, Shalem O, Lee K, Shi X, Scott DA, Song J, Pan JQ, Weissleder R, Lee H, Zhang F, Sharp PA. Cell. 2015 Mar 12;160(6):1246-60. doi: 10.1016/j.cell.2015.02.038. Epub 2015 Mar 5. 10.1016/j.cell.2015.02.038 PubMed 25748654 |
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 lentiCas9-EGFP 13177 bp ds-DNA circular SYN 18-JAN-2019
DEFINITION Expresses human codon-optimized S. pyogenes Cas9 protein and EGFP
from EFS promoter. 3rd generation lentiviral backbone. .
ACCESSION .
VERSION .
KEYWORDS lentiCas9-EGFP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 13177)
AUTHORS Chen S, Sanjana NE, Zheng K, Shalem O, Lee K, Shi X, Scott DA, Song
J, Pan JQ, Weissleder R, Lee H, Zhang F, Sharp PA
TITLE Genome-wide CRISPR screen in a mouse model of tumor growth and
metastasis.
JOURNAL Cell. 2015 Mar 12;160(6):1246-60. doi: 10.1016/j.cell.2015.02.038.
Epub 2015 Mar 5.
PUBMED 25748654
REFERENCE 2 (bases 1 to 13177)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Jan 18, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..13177
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(47..66)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 238..617
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 619..817
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 772..792
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
LTR 835..1015
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 1062..1187
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1680..1913
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
misc_feature 2440..2557
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2650..2861
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
regulatory 2880..2889
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2886..6989
/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"
CDS 6990..7037
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS 7038..7061
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
CDS 7071..7127
/codon_start=1
/product="2A peptide from porcine teschovirus-1
polyprotein"
/label=P2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="ATNFSLLKQAGDVEENPGP"
CDS 7128..7844
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/note="mammalian codon-optimized"
/translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKA
NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
FVTAAGITLGMDELYK"
primer_bind complement(7170..7191)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(7431..7450)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 7778..7799
/label=EGFP-C
/note="EGFP, forward primer"
misc_feature 7869..8457
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(7922..7942)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(8340..8351)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(8460..8476)
/label=KS primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(8461..8477)
/label=pBluescriptKS
/note="For pBluescript vector"
LTR 8982..9162
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
primer_bind complement(9188..9205)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 9194..9418
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 9464..9892
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(9551..9570)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 9761..9782
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(9901..9921)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 9906..10235
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 10086..10221
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 10148..10167
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
promoter 10283..10330
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 10349..10723
/codon_start=1
/gene="Sh ble from Streptoalloteichus hindustanus"
/product="antibiotic-binding protein"
/label=BleoR
/note="confers resistance to bleomycin, phleomycin, and
Zeocin(TM)"
/translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
EFALRDPAGNCVHFVAEEQD"
polyA_signal 10853..10974
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(10890..10909)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 10944..10963
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(11023..11039)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(11023..11039)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(11036..11058)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 11047..11063
/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(11071..11101)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 11116..11137
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(11254..11271)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(11425..12013)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(11505..11524)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(12184..13044)
/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 12807..12826
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(13045..13149)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
2581 agatccagtt tggttaatta gctagctagg tcttgaaagg agtgggaatt ggctccggtg
2641 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
2701 gcaattgatc cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
2761 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
2821 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggaccggttc tagagcgctg
2881 ccaccatgga caagaagtac agcatcggcc tggacatcgg caccaactct gtgggctggg
2941 ccgtgatcac cgacgagtac aaggtgccca gcaagaaatt caaggtgctg ggcaacaccg
3001 accggcacag catcaagaag aacctgatcg gagccctgct gttcgacagc ggcgaaacag
3061 ccgaggccac ccggctgaag agaaccgcca gaagaagata caccagacgg aagaaccgga
3121 tctgctatct gcaagagatc ttcagcaacg agatggccaa ggtggacgac agcttcttcc
3181 acagactgga agagtccttc ctggtggaag aggataagaa gcacgagcgg caccccatct
3241 tcggcaacat cgtggacgag gtggcctacc acgagaagta ccccaccatc taccacctga
3301 gaaagaaact ggtggacagc accgacaagg ccgacctgcg gctgatctat ctggccctgg
3361 cccacatgat caagttccgg ggccacttcc tgatcgaggg cgacctgaac cccgacaaca
3421 gcgacgtgga caagctgttc atccagctgg tgcagaccta caaccagctg ttcgaggaaa
3481 accccatcaa cgccagcggc gtggacgcca aggccatcct gtctgccaga ctgagcaaga
3541 gcagacggct ggaaaatctg atcgcccagc tgcccggcga gaagaagaat ggcctgttcg
3601 gaaacctgat tgccctgagc ctgggcctga cccccaactt caagagcaac ttcgacctgg
3661 ccgaggatgc caaactgcag ctgagcaagg acacctacga cgacgacctg gacaacctgc
3721 tggcccagat cggcgaccag tacgccgacc tgtttctggc cgccaagaac ctgtccgacg
3781 ccatcctgct gagcgacatc ctgagagtga acaccgagat caccaaggcc cccctgagcg
3841 cctctatgat caagagatac gacgagcacc accaggacct gaccctgctg aaagctctcg
3901 tgcggcagca gctgcctgag aagtacaaag agattttctt cgaccagagc aagaacggct
3961 acgccggcta cattgacggc ggagccagcc aggaagagtt ctacaagttc atcaagccca
4021 tcctggaaaa gatggacggc accgaggaac tgctcgtgaa gctgaacaga gaggacctgc
4081 tgcggaagca gcggaccttc gacaacggca gcatccccca ccagatccac ctgggagagc
4141 tgcacgccat tctgcggcgg caggaagatt tttacccatt cctgaaggac aaccgggaaa
4201 agatcgagaa gatcctgacc ttccgcatcc cctactacgt gggccctctg gccaggggaa
4261 acagcagatt cgcctggatg accagaaaga gcgaggaaac catcaccccc tggaacttcg
4321 aggaagtggt ggacaagggc gcttccgccc agagcttcat cgagcggatg accaacttcg
4381 ataagaacct gcccaacgag aaggtgctgc ccaagcacag cctgctgtac gagtacttca
4441 ccgtgtataa cgagctgacc aaagtgaaat acgtgaccga gggaatgaga aagcccgcct
4501 tcctgagcgg cgagcagaaa aaggccatcg tggacctgct gttcaagacc aaccggaaag
4561 tgaccgtgaa gcagctgaaa gaggactact tcaagaaaat cgagtgcttc gactccgtgg
4621 aaatctccgg cgtggaagat cggttcaacg cctccctggg cacataccac gatctgctga
4681 aaattatcaa ggacaaggac ttcctggaca atgaggaaaa cgaggacatt ctggaagata
4741 tcgtgctgac cctgacactg tttgaggaca gagagatgat cgaggaacgg ctgaaaacct
4801 atgcccacct gttcgacgac aaagtgatga agcagctgaa gcggcggaga tacaccggct
4861 ggggcaggct gagccggaag ctgatcaacg gcatccggga caagcagtcc ggcaagacaa
4921 tcctggattt cctgaagtcc gacggcttcg ccaacagaaa cttcatgcag ctgatccacg
4981 acgacagcct gacctttaaa gaggacatcc agaaagccca ggtgtccggc cagggcgata
5041 gcctgcacga gcacattgcc aatctggccg gcagccccgc cattaagaag ggcatcctgc
5101 agacagtgaa ggtggtggac gagctcgtga aagtgatggg ccggcacaag cccgagaaca
5161 tcgtgatcga aatggccaga gagaaccaga ccacccagaa gggacagaag aacagccgcg
5221 agagaatgaa gcggatcgaa gagggcatca aagagctggg cagccagatc ctgaaagaac
5281 accccgtgga aaacacccag ctgcagaacg agaagctgta cctgtactac ctgcagaatg
5341 ggcgggatat gtacgtggac caggaactgg acatcaaccg gctgtccgac tacgatgtgg
5401 accatatcgt gcctcagagc tttctgaagg acgactccat cgacaacaag gtgctgacca
5461 gaagcgacaa gaaccggggc aagagcgaca acgtgccctc cgaagaggtc gtgaagaaga
5521 tgaagaacta ctggcggcag ctgctgaacg ccaagctgat tacccagaga aagttcgaca
5581 atctgaccaa ggccgagaga ggcggcctga gcgaactgga taaggccggc ttcatcaaga
5641 gacagctggt ggaaacccgg cagatcacaa agcacgtggc acagatcctg gactcccgga
5701 tgaacactaa gtacgacgag aatgacaagc tgatccggga agtgaaagtg atcaccctga
5761 agtccaagct ggtgtccgat ttccggaagg atttccagtt ttacaaagtg cgcgagatca
5821 acaactacca ccacgcccac gacgcctacc tgaacgccgt cgtgggaacc gccctgatca
5881 aaaagtaccc taagctggaa agcgagttcg tgtacggcga ctacaaggtg tacgacgtgc
5941 ggaagatgat cgccaagagc gagcaggaaa tcggcaaggc taccgccaag tacttcttct
6001 acagcaacat catgaacttt ttcaagaccg agattaccct ggccaacggc gagatccgga
6061 agcggcctct gatcgagaca aacggcgaaa ccggggagat cgtgtgggat aagggccggg
6121 attttgccac cgtgcggaaa gtgctgagca tgccccaagt gaatatcgtg aaaaagaccg
6181 aggtgcagac aggcggcttc agcaaagagt ctatcctgcc caagaggaac agcgataagc
6241 tgatcgccag aaagaaggac tgggacccta agaagtacgg cggcttcgac agccccaccg
6301 tggcctattc tgtgctggtg gtggccaaag tggaaaaggg caagtccaag aaactgaaga
6361 gtgtgaaaga gctgctgggg atcaccatca tggaaagaag cagcttcgag aagaatccca
6421 tcgactttct ggaagccaag ggctacaaag aagtgaaaaa ggacctgatc atcaagctgc
6481 ctaagtactc cctgttcgag ctggaaaacg gccggaagag aatgctggcc tctgccggcg
6541 aactgcagaa gggaaacgaa ctggccctgc cctccaaata tgtgaacttc ctgtacctgg
6601 ccagccacta tgagaagctg aagggctccc ccgaggataa tgagcagaaa cagctgtttg
6661 tggaacagca caagcactac ctggacgaga tcatcgagca gatcagcgag ttctccaaga
6721 gagtgatcct ggccgacgct aatctggaca aagtgctgtc cgcctacaac aagcaccggg
6781 ataagcccat cagagagcag gccgagaata tcatccacct gtttaccctg accaatctgg
6841 gagcccctgc cgccttcaag tactttgaca ccaccatcga ccggaagagg tacaccagca
6901 ccaaagaggt gctggacgcc accctgatcc accagagcat caccggcctg tacgagacac
6961 ggatcgacct gtctcagctg ggaggcgaca agcgacctgc cgccacaaag aaggctggac
7021 aggctaagaa gaagaaagat tacaaagacg atgacgataa gggatccggc gcaacaaact
7081 tctctctgct gaaacaagcc ggagatgtcg aagagaatcc tggaccggtg agcaagggcg
7141 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
7201 acaagttcag cgtgtctggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
7261 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
7321 cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca
7381 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
7441 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
7501 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
7561 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggcgaact
7621 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
7681 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
7741 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
7801 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaagaattc gatatcaagc
7861 ttatcggtaa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
7921 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
7981 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
8041 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
8101 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
8161 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
8221 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
8281 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
8341 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
8401 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
8461 gataccgtcg acctcgagac ctagaaaaac atggagcaat cacaagtagc aatacagcag
8521 ctaccaatgc tgattgtgcc tggctagaag cacaagagga ggaggaggtg ggttttccag
8581 tcacacctca ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact
8641 ttttaaaaga aaagggggga ctggaagggc taattcactc ccaacgaaga caagatatcc
8701 ttgatctgtg gatctaccac acacaaggct acttccctga ttggcagaac tacacaccag
8761 ggccagggat cagatatcca ctgacctttg gatggtgcta caagctagta ccagttgagc
8821 aagagaaggt agaagaagcc aatgaaggag agaacacccg cttgttacac cctgtgagcc
8881 tgcatgggat ggatgacccg gagagagaag tattagagtg gaggtttgac agccgcctag
8941 catttcatca catggcccga gagctgcatc cggactgtac tgggtctctc tggttagacc
9001 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
9061 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
9121 gatccctcag acccttttag tcagtgtgga aaatctctag cagggcccgt ttaaacccgc
9181 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
9241 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
9301 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
9361 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
9421 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
9481 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
9541 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
9601 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
9661 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
9721 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
9781 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
9841 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
9901 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
9961 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
10021 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
10081 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
10141 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
10201 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
10261 ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt
10321 ataatacgac aaggtgagga actaaaccat ggccaagttg accagtgccg ttccggtgct
10381 caccgcgcgc gacgtcgccg gagcggtcga gttctggacc gaccggctcg ggttctcccg
10441 ggacttcgtg gaggacgact tcgccggtgt ggtccgggac gacgtgaccc tgttcatcag
10501 cgcggtccag gaccaggtgg tgccggacaa caccctggcc tgggtgtggg tgcgcggcct
10561 ggacgagctg tacgccgagt ggtcggaggt cgtgtccacg aacttccggg acgcctccgg
10621 gccggccatg accgagatcg gcgagcagcc gtgggggcgg gagttcgccc tgcgcgaccc
10681 ggccggcaac tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc tacgagattt
10741 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
10801 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
10861 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
10921 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
10981 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
11041 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
11101 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
11161 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
11221 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
11281 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
11341 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
11401 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
11461 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
11521 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
11581 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
11641 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
11701 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
11761 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
11821 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
11881 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
11941 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
12001 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
12061 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
12121 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
12181 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
12241 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
12301 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
12361 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
12421 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
12481 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
12541 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
12601 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
12661 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
12721 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
12781 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
12841 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
12901 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
12961 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
13021 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
13081 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
13141 ggttccgcgc acatttcccc gaaaagtgcc acctgac
//
