| 质粒名称: | pFUGW SpCas9 |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 131506 |
| 存储实验室: | Harold MacGillavry |
|
载体骨架基本信息 |
|
| 载体名称: | pFUGW |
| 完整质粒大小: | 13541bp |
| 载体类型: | 哺乳细胞表达 |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | SpCas9 |
| 物种来源: | S. pyogenes |
| 插入大小: | 951 bp |
| 基因突变: | T2A, S13L, A92T, V104T, V114I, E161K, Y193K, F198Y, D202T, I203V, Y258F, A283V, K288T, N290Y relative to RpBphP2 |
| GenBank编号 | JN247409 |
| 启动子: | hUBC |
| 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: | 未知(未破坏) |
| 3’克隆位点: | 未知(未破坏) |
| 5’测序引物 | GGCGAGTGTGTTTTGTGAAG |
| 3’测序引物 | GCAGCGTATCCACATAGCG |
|
文献引用方法 |
|
| 材料和方法部分: | pFUGW SpCas9 was a gift from Harold MacGillavry (Addgene plasmid # 131506 ; http://n2t.net/addgene:131506 ; RRID:Addgene_131506) |
| 参考文献部分: | ORANGE: A CRISPR/Cas9-based genome editing toolbox for epitope tagging of endogenous proteins in neurons. Willems J, de Jong APH, Scheefhals N, Mertens E, Catsburg LAE, Poorthuis RB, de Winter F, Verhaagen J, Meye FJ, MacGillavry HD. PLoS Biol. 2020 Apr 10;18(4):e3000665. doi: 10.1371/journal.pbio.3000665. 10.1371/journal.pbio.3000665 PubM |
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 pFUGW SpCas9 13418 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION lentiviral expression of SpCas9 under hUBC promotor.
ACCESSION .
VERSION .
KEYWORDS pFUGW SpCas9
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 13418)
AUTHORS Willems J, de Jong APH, Scheefhals N, Mertens E, Catsburg LAE,
Poorthuis RB, de Winter F, Verhaagen J, Meye FJ, MacGillavry HD
TITLE ORANGE: A CRISPR/Cas9-based genome editing toolbox for epitope
tagging of endogenous proteins in neurons.
JOURNAL PLoS Biol. 2020 Apr 10;18(4):e3000665. doi:
10.1371/journal.pbio.3000665.
PUBMED 32275651
REFERENCE 2 (bases 1 to 13418)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..13418
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 262..281
/label=hUBCpro-F
/note="Human Ubiquitin C (UbC) promoter, forward primer"
CDS 481..507
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
CDS 511..531
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 541..4641
/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 4645..4665
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
misc_feature 4702..5290
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(4755..4775)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(5173..5184)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(5293..5309)
/label=KS primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5294..5310)
/label=pBluescriptKS
/note="For pBluescript vector"
LTR 5815..5995
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
primer_bind complement(6021..6038)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 6027..6251
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 6297..6725
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(6384..6403)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 6594..6615
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(6734..6754)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 6739..7068
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 6919..7054
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 6981..7000
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
promoter 7116..7163
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 7182..7556
/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 7686..7807
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(7723..7742)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 7777..7796
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(7856..7872)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(7856..7872)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(7869..7891)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 7880..7896
/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(7904..7934)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 7949..7970
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(8087..8104)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(8258..8846)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(8338..8357)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(9017..9877)
/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 9640..9659
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(9878..9982)
/gene="bla"
/label=AmpR promoter
primer_bind complement(10057..10076)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 10248..10627
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 10629..10827
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 10782..10802
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
LTR 10845..11025
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 11072..11197
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 11690..11923
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 12108..12152
/codon_start=1
/product="antigenic peptide corresponding to amino acids
655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
et al., 2013)"
/label=gp41 peptide
/note="recognized by the 2H10 single-chain llama nanobody"
/translation="KNEQELLELDKWASL"
misc_feature 12450..12567
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 12632..425
/label=UbC promoter
/note="human ubiquitin C promoter"
ORIGIN
1 gtgccgttgg gcagtgcacc cgtacctttg ggagcgcgcg ccctcgtcgt gtcgtgacgt
61 cacccgttct gttggcttat aatgcagggt ggggccacct gccggtaggt gtgcggtagg
121 cttttctccg tcgcaggacg cagggttcgg gcctagggta ggctctcctg aatcgacagg
181 cgccggacct ctggtgaggg gagggataag tgaggcgtca gtttctttgg tcggttttat
241 gtacctatct tcttaagtag ctgaagctcc ggttttgaac tatgcgctcg gggttggcga
301 gtgtgttttg tgaagttttt taggcacctt ttgaaatgta atcatttggg tcaatatgta
361 attttcagtg ttagactagt aaattgtccg ctaaattctg gccgtttttg gcttttttgt
421 tagacgaagc ttgggctgca ggtcgactct agaggatccc cgggtaccgg tgccaccatg
481 tacccatacg atgttccaga ttacgcttcg ccgaagaaaa agcgcaaggt cgaagcgtcc
541 gacaagaagt acagcatcgg cctggacatc ggcaccaact ctgtgggctg ggccgtgatc
601 accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac
661 agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc
721 acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat
781 ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg
841 gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac
901 atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa
961 ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg
1021 atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg
1081 gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc
1141 aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg
1201 ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg
1261 attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat
1321 gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag
1381 atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg
1441 ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg
1501 atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag
1561 cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc
1621 tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa
1681 aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag
1741 cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc
1801 attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag
1861 aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga
1921 ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg
1981 gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac
2041 ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat
2101 aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc
2161 ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg
2221 aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc
2281 ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc
2341 aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg
2401 accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac
2461 ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg
2521 ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat
2581 ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc
2641 ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac
2701 gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg
2761 aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc
2821 gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg
2881 aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg
2941 gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat
3001 atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc
3061 gtgcctcaga gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac
3121 aagaaccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac
3181 tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc
3241 aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg
3301 gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact
3361 aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag
3421 ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac
3481 caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac
3541 cctaagctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg
3601 atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac
3661 atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct
3721 ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc
3781 accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag
3841 acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc
3901 agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat
3961 tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa
4021 gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt
4081 ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac
4141 tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag
4201 aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac
4261 tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag
4321 cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc
4381 ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc
4441 atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct
4501 gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag
4561 gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac
4621 ctgtctcagc tgggaggcga cagccccaag aagaagagaa aggtggaggc cagctaagaa
4681 ttcgatatca agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact
4741 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
4801 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg
4861 ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg
4921 tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct cctttccggg
4981 actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc
5041 tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca
5101 tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc
5161 tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct
5221 ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc
5281 gcctccccgc atcgataccg tcgacctcga gacctagaaa aacatggagc aatcacaagt
5341 agcaatacag cagctaccaa tgctgattgt gcctggctag aagcacaaga ggaggaggag
5401 gtgggttttc cagtcacacc tcaggtacct ttaagaccaa tgacttacaa ggcagctgta
5461 gatcttagcc actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga
5521 agacaagata tccttgatct gtggatctac cacacacaag gctacttccc tgattggcag
5581 aactacacac cagggccagg gatcagatat ccactgacct ttggatggtg ctacaagcta
5641 gtaccagttg agcaagagaa ggtagaagaa gccaatgaag gagagaacac ccgcttgtta
5701 caccctgtga gcctgcatgg gatggatgac ccggagagag aagtattaga gtggaggttt
5761 gacagccgcc tagcatttca tcacatggcc cgagagctgc atccggactg tactgggtct
5821 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
5881 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
5941 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagggcc
6001 cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
6061 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
6121 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
6181 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
6241 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
6301 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
6361 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
6421 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
6481 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
6541 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
6601 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
6661 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
6721 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
6781 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
6841 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
6901 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
6961 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
7021 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
7081 gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt
7141 atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag ttgaccagtg
7201 ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc
7261 tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga
7321 ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt
7381 gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc
7441 gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg
7501 ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg
7561 tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
7621 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
7681 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
7741 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
7801 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
7861 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
7921 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
7981 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
8041 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
8101 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
8161 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
8221 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
8281 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
8341 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
8401 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
8461 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
8521 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
8581 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
8641 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
8701 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
8761 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
8821 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
8881 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
8941 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
9001 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
9061 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
9121 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
9181 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
9241 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
9301 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
9361 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
9421 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
9481 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
9541 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
9601 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
9661 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
9721 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
9781 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
9841 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
9901 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
9961 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
10021 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
10081 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
10141 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
10201 taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac attgattatt
10261 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
10321 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
10381 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
10441 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
10501 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
10561 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
10621 taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
10681 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
10741 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
10801 tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct ctctggttag
10861 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
10921 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
10981 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag
11041 ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga
11101 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
11161 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
11221 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
11281 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
11341 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
11401 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
11461 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
11521 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
11581 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
11641 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
11701 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
11761 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
11821 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
11881 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
11941 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
12001 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
12061 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
12121 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
12181 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
12241 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
12301 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag
12361 agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt gcgccaattc
12421 tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat tggggggtac
12481 agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa agaattacaa
12541 aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag agatccagtt
12601 tggttaatta acccgtgtcg gctccagatc tggcctccgc gccgggtttt ggcgcctccc
12661 gcgggcgccc ccctcctcac ggcgagcgct gccacgtcag acgaagggcg cagcgagcgt
12721 cctgatcctt ccgcccggac gctcaggaca gcggcccgct gctcataaga ctcggcctta
12781 gaaccccagt atcagcagaa ggacatttta ggacgggact tgggtgactc tagggcactg
12841 gttttctttc cagagagcgg aacaggcgag gaaaagtagt cccttctcgg cgattctgcg
12901 gagggatctc cgtggggcgg tgaacgccga tgattatata aggacgcgcc gggtgtggca
12961 cagctagttc cgtcgcagcc gggatttggg tcgcggttct tgtttgtgga tcgctgtgat
13021 cgtcacttgg tgagtagcgg gctgctgggc tggccggggc tttcgtggcc gccgggccgc
13081 tcggtgggac ggaagcgtgt ggagagaccg ccaagggctg tagtctgggt ccgcgagcaa
13141 ggttgccctg aactgggggt tggggggagc gcagcaaaat ggcggctgtt cccgagtctt
13201 gaatggaaga cgcttgtgag gcgggctgtg aggtcgttga aacaaggtgg ggggcatggt
13261 gggcggcaag aacccaaggt cttgaggcct tcgctaatgc gggaaagctc ttattcgggt
13321 gagatgggct ggggcaccat ctggggaccc tgacgtgaag tttgtcactg actggagaac
13381 tcggtttgtc gtctgttgcg ggggcggcag ttatggcg
//
