| 质粒名称: | lentiCRISPR - EGFP sgRNA 1 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 51760 |
| 存储实验室: | Feng Zhang |
|
载体骨架基本信息 |
|
| 载体名称: | lentiCRISPR (pXPR_001), Addgene plasmid #49535 |
| 载体出品公司: | Zhang lab |
| 载体抗性: | 氨苄青霉素 |
| 载体类型: | 哺乳动物表达，慢病毒，CRISPR |
| 筛选标记: | 嘌呤霉素 |
| 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,S. pyogenes CRISPR-Cas9 |
| 物种来源: | 合成的 |
| 插入大小: | 4200 bp |
| 启动子 | EFS |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | BamHI(未破坏) |
| 3' 克隆位点: | NheI(未破坏) |
| 5' 测序引物 | GGTACAGTGCAGGGGAAAGA |
| 3' 测序引物 | TGCCCTCCAAATATGTGAACT |
|
文献引用方法 |
|
| 材料和方法部分: | lentiCRISPR - EGFP sgRNA 1 was a gift from Feng Zhang (Addgene plasmid # 51760 ; http://n2t.net/addgene:51760 ; RRID:Addgene_51760) |
| 参考文献部分: | Genome-scale CRISPR-Cas9 knockout screening in human cells. Shalem O, Sanjana NE, Hartenian E, Shi X, Scott DA, Mikkelsen TS, Heckl D, Ebert BL, Root DE, Doench JG, Zhang F. Science. 2014 Jan 3;343(6166):84-7. doi: 10.1126/science.1247005. Epub 2013 Dec 12. 10.1126/science.1247005 PubMed 24336571 |
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 lentiCRISPR - EGFP sgRNA 1 11589 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Expresses human codon-optimized Cas9 protein and puromycin
resistance from EFS promoter and an EGFP targeting synthetic
single-guide RNA (sgRNA) element from U6 promoter. Lentiviral
backbone..
ACCESSION .
VERSION .
KEYWORDS lentiCRISPR - EGFP sgRNA 1
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 11589)
AUTHORS Shalem O, Sanjana NE, Hartenian E, Shi X, Scott DA, Mikkelsen TS,
Heckl D, Ebert BL, Root DE, Doench JG, Zhang F
TITLE Genome-scale CRISPR-Cas9 knockout screening in human cells.
JOURNAL Science. 2014 Jan 3;343(6166):84-7. doi: 10.1126/science.1247005.
Epub 2013 Dec 12.
PUBMED 24336571
REFERENCE 2 (bases 1 to 11589)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..11589
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 5..231
/label=RSV promoter
/note="Rous sarcoma virus enhancer/promoter"
LTR 232..412
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 459..584
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1077..1310
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
promoter 1717..1957
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 1717..1737
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 1888..1907
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 1986..2061
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
misc_feature 2109..2226
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2295..2506
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
regulatory 2526..2535
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2535..2558
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
CDS 2565..2585
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 2610..6710
/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
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 6711..6758
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS 6774..6830
/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 6831..7430
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind complement(6831..6850)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
primer_bind 7327..7347
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
misc_feature 7446..8034
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(7499..7519)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(7917..7928)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
LTR 8106..8339
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 8417..8538
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(8454..8473)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 8508..8527
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
rep_origin 8578..8713
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 8640..8659
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind complement(8733..8752)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(8734..8752)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(8762..8779)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(8762..8778)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(8771..8793)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 8920..9375
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(9007..9026)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 9217..9238
/label=F1ori-F
/note="F1 origin, forward primer"
promoter 9401..9505
/gene="bla"
/label=AmpR promoter
CDS 9506..10366
/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 complement(9724..9743)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 10537..11125
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 11026..11045
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 11279..11296
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 11413..11434
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 11449..11479
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 11487..11503
/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)."
primer_bind 11492..11514
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 11511..11527
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 11511..11527
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind 11546..11566
/label=T3
/note="T3 promoter, forward primer"
promoter 11548..11566
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
1 ttaatgtagt cttatgcaat actcttgtag tcttgcaaca tggtaacgat gagttagcaa
61 catgccttac aaggagagaa aaagcaccgt gcatgccgat tggtggaagt aaggtggtac
121 gatcgtgcct tattaggaag gcaacagacg ggtctgacat ggattggacg aaccactgaa
181 ttgccgcatt gcagagatat tgtatttaag tgcctagctc gatacataaa cgggtctctc
241 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
301 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
361 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
421 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
481 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
541 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
601 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
661 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
721 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
781 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
841 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
901 gtaagaccac cgcacagcaa gcggccgctg atcttcagac ctggaggagg agatatgagg
961 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
1021 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
1081 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
1141 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
1201 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
1261 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
1321 ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag ttggagtaat
1381 aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag agaaattaac
1441 aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca agaaaagaat
1501 gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt taacataaca
1561 aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt aggtttaaga
1621 atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc accattatcg
1681 tttcagaccc acctcccaac cccgagggga cccagagagg gcctatttcc catgattcct
1741 tcatatttgc atatacgata caaggctgtt agagagataa ttagaattaa tttgactgta
1801 aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt
1861 gcagttttaa aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat
1921 ttcgatttct tggctttata tatcttgtgg aaaggacgaa acaccgggcg aggagctgtt
1981 caccggtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa
2041 aaagtggcac cgagtcggtg cttttttgaa ttctagatct tgagacaaat ggcagtattc
2101 atccacaatt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga aagaatagta
2161 gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac aaaaattcaa
2221 aattttcggg tttattacag ggacagcaga gatccacttt ggcgccggct cgagtggctc
2281 cggtgcccgt cagtgggcag agcgcacatc gcccacagtc cccgagaagt tggggggagg
2341 ggtcggcaat tgaaccggtg cctagagaag gtggcgcggg gtaaactggg aaagtgatgt
2401 cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt
2461 cgccgtgaac gttctttttc gcaacgggtt tgccgccaga acacaggtgt cgtgacgcgg
2521 gatccgccac catggattac aaagacgatg acgataagat ggccccaaag aagaagcgga
2581 aggtcggtat ccacggagtc ccagcagccg acaagaagta cagcatcggc ctggacatcg
2641 gcaccaactc tgtgggctgg gccgtgatca ccgacgagta caaggtgccc agcaagaaat
2701 tcaaggtgct gggcaacacc gaccggcaca gcatcaagaa gaacctgatc ggagccctgc
2761 tgttcgacag cggcgaaaca gccgaggcca cccggctgaa gagaaccgcc agaagaagat
2821 acaccagacg gaagaaccgg atctgctatc tgcaagagat cttcagcaac gagatggcca
2881 aggtggacga cagcttcttc cacagactgg aagagtcctt cctggtggaa gaggataaga
2941 agcacgagcg gcaccccatc ttcggcaaca tcgtggacga ggtggcctac cacgagaagt
3001 accccaccat ctaccacctg agaaagaaac tggtggacag caccgacaag gccgacctgc
3061 ggctgatcta tctggccctg gcccacatga tcaagttccg gggccacttc ctgatcgagg
3121 gcgacctgaa ccccgacaac agcgacgtgg acaagctgtt catccagctg gtgcagacct
3181 acaaccagct gttcgaggaa aaccccatca acgccagcgg cgtggacgcc aaggccatcc
3241 tgtctgccag actgagcaag agcagacggc tggaaaatct gatcgcccag ctgcccggcg
3301 agaagaagaa tggcctgttc ggcaacctga ttgccctgag cctgggcctg acccccaact
3361 tcaagagcaa cttcgacctg gccgaggatg ccaaactgca gctgagcaag gacacctacg
3421 acgacgacct ggacaacctg ctggcccaga tcggcgacca gtacgccgac ctgtttctgg
3481 ccgccaagaa cctgtccgac gccatcctgc tgagcgacat cctgagagtg aacaccgaga
3541 tcaccaaggc ccccctgagc gcctctatga tcaagagata cgacgagcac caccaggacc
3601 tgaccctgct gaaagctctc gtgcggcagc agctgcctga gaagtacaaa gagattttct
3661 tcgaccagag caagaacggc tacgccggct acattgacgg cggagccagc caggaagagt
3721 tctacaagtt catcaagccc atcctggaaa agatggacgg caccgaggaa ctgctcgtga
3781 agctgaacag agaggacctg ctgcggaagc agcggacctt cgacaacggc agcatccccc
3841 accagatcca cctgggagag ctgcacgcca ttctgcggcg gcaggaagat ttttacccat
3901 tcctgaagga caaccgggaa aagatcgaga agatcctgac cttccgcatc ccctactacg
3961 tgggccctct ggccagggga aacagcagat tcgcctggat gaccagaaag agcgaggaaa
4021 ccatcacccc ctggaacttc gaggaagtgg tggacaaggg cgcttccgcc cagagcttca
4081 tcgagcggat gaccaacttc gataagaacc tgcccaacga gaaggtgctg cccaagcaca
4141 gcctgctgta cgagtacttc accgtgtata acgagctgac caaagtgaaa tacgtgaccg
4201 agggaatgag aaagcccgcc ttcctgagcg gcgagcagaa aaaggccatc gtggacctgc
4261 tgttcaagac caaccggaaa gtgaccgtga agcagctgaa agaggactac ttcaagaaaa
4321 tcgagtgctt cgactccgtg gaaatctccg gcgtggaaga tcggttcaac gcctccctgg
4381 gcacatacca cgatctgctg aaaattatca aggacaagga cttcctggac aatgaggaaa
4441 acgaggacat tctggaagat atcgtgctga ccctgacact gtttgaggac agagagatga
4501 tcgaggaacg gctgaaaacc tatgcccacc tgttcgacga caaagtgatg aagcagctga
4561 agcggcggag atacaccggc tggggcaggc tgagccggaa gctgatcaac ggcatccggg
4621 acaagcagtc cggcaagaca atcctggatt tcctgaagtc cgacggcttc gccaacagaa
4681 acttcatgca gctgatccac gacgacagcc tgacctttaa agaggacatc cagaaagccc
4741 aggtgtccgg ccagggcgat agcctgcacg agcacattgc caatctggcc ggcagccccg
4801 ccattaagaa gggcatcctg cagacagtga aggtggtgga cgagctcgtg aaagtgatgg
4861 gccggcacaa gcccgagaac atcgtgatcg aaatggccag agagaaccag accacccaga
4921 agggacagaa gaacagccgc gagagaatga agcggatcga agagggcatc aaagagctgg
4981 gcagccagat cctgaaagaa caccccgtgg aaaacaccca gctgcagaac gagaagctgt
5041 acctgtacta cctgcagaat gggcgggata tgtacgtgga ccaggaactg gacatcaacc
5101 ggctgtccga ctacgatgtg gaccatatcg tgcctcagag ctttctgaag gacgactcca
5161 tcgacaacaa ggtgctgacc agaagcgaca agaaccgggg caagagcgac aacgtgccct
5221 ccgaagaggt cgtgaagaag atgaagaact actggcggca gctgctgaac gccaagctga
5281 ttacccagag aaagttcgac aatctgacca aggccgagag aggcggcctg agcgaactgg
5341 ataaggccgg cttcatcaag agacagctgg tggaaacccg gcagatcaca aagcacgtgg
5401 cacagatcct ggactcccgg atgaacacta agtacgacga gaatgacaag ctgatccggg
5461 aagtgaaagt gatcaccctg aagtccaagc tggtgtccga tttccggaag gatttccagt
5521 tttacaaagt gcgcgagatc aacaactacc accacgccca cgacgcctac ctgaacgccg
5581 tcgtgggaac cgccctgatc aaaaagtacc ctaagctgga aagcgagttc gtgtacggcg
5641 actacaaggt gtacgacgtg cggaagatga tcgccaagag cgagcaggaa atcggcaagg
5701 ctaccgccaa gtacttcttc tacagcaaca tcatgaactt tttcaagacc gagattaccc
5761 tggccaacgg cgagatccgg aagcggcctc tgatcgagac aaacggcgaa accggggaga
5821 tcgtgtggga taagggccgg gattttgcca ccgtgcggaa agtgctgagc atgccccaag
5881 tgaatatcgt gaaaaagacc gaggtgcaga caggcggctt cagcaaagag tctatcctgc
5941 ccaagaggaa cagcgataag ctgatcgcca gaaagaagga ctgggaccct aagaagtacg
6001 gcggcttcga cagccccacc gtggcctatt ctgtgctggt ggtggccaaa gtggaaaagg
6061 gcaagtccaa gaaactgaag agtgtgaaag agctgctggg gatcaccatc atggaaagaa
6121 gcagcttcga gaagaatccc atcgactttc tggaagccaa gggctacaaa gaagtgaaaa
6181 aggacctgat catcaagctg cctaagtact ccctgttcga gctggaaaac ggccggaaga
6241 gaatgctggc ctctgccggc gaactgcaga agggaaacga actggccctg ccctccaaat
6301 atgtgaactt cctgtacctg gccagccact atgagaagct gaagggctcc cccgaggata
6361 atgagcagaa acagctgttt gtggaacagc acaagcacta cctggacgag atcatcgagc
6421 agatcagcga gttctccaag agagtgatcc tggccgacgc taatctggac aaagtgctgt
6481 ccgcctacaa caagcaccgg gataagccca tcagagagca ggccgagaat atcatccacc
6541 tgtttaccct gaccaatctg ggagcccctg ccgccttcaa gtactttgac accaccatcg
6601 accggaagag gtacaccagc accaaagagg tgctggacgc caccctgatc caccagagca
6661 tcaccggcct gtacgagaca cggatcgacc tgtctcagct gggaggcgac aagcgtcctg
6721 ctgctactaa gaaagctggt caagctaaga aaaagaaagc tagcggcagc ggcgccacca
6781 acttcagcct gctgaagcag gccggcgacg tggaggagaa ccccggcccc atgaccgagt
6841 acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg
6901 ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg
6961 agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg
7021 tgtgggtcgc ggacgacggc gcggccgtgg cggtctggac cacgccggag agcgtcgaag
7081 cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg
7141 ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt
7201 tcctggccac cgtcggagtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg
7261 tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg
7321 cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg
7381 tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga acgcgttaag
7441 tcgacaatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
7501 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
7561 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
7621 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
7681 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
7741 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
7801 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
7861 tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
7921 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
7981 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcgtcgac
8041 tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg
8101 gggactggaa gggctaattc actcccaacg aagacaagat ctgctttttg cttgtactgg
8161 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
8221 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
8281 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
8341 tacgtatagt agttcatgtc atcttattat tcagtattta taacttgcaa agaaatgaat
8401 atcagagagt gagaggaact tgtttattgc agcttataat ggttacaaat aaagcaatag
8461 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
8521 actcatcaat gtatcttatc atgtctggct ctagctatcc cgcccctaac tccgcccatc
8581 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
8641 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
8701 ttttttggag gcctagggac gtacccaatt cgccctatag tgagtcgtat tacgcgcgct
8761 cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc
8821 gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc
8881 gcccttccca acagttgcgc agcctgaatg gcgaatggga cgcgccctgt agcggcgcat
8941 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
9001 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
9061 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
9121 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
9181 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
9241 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
9301 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
9361 taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
9421 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
9481 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
9541 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
9601 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
9661 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
9721 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
9781 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
9841 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
9901 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
9961 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
10021 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
10081 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
10141 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
10201 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
10261 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
10321 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
10381 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
10441 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
10501 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
10561 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
10621 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
10681 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
10741 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
10801 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
10861 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
10921 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
10981 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
11041 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
11101 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
11161 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
11221 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
11281 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
11341 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
11401 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
11461 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
11521 ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctgga
11581 gctgcaagc
//
