| 质粒名称: | pET-PE2-His (IK1822) |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 170103 |
| 存储实验室: | Keith Joung |
|
载体骨架基本信息 |
|
| 载体名称: | pET |
| 载体出品公司: | Clontech |
| 空载体大小: | 5238bp |
| 完整质粒大小: | 11619bp |
| 载体类型: | Bacterial Expression, CRISPR |
| 载体抗性: | Kanamycin |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 低拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | bpNLS-nSpCas9(H840A)-MMLVRT-bpNLS-6xHis |
| 物种来源: | Synthetic; Streptococcus pyogenes and Moloney Murine Leukemia Virus |
| 插入大小: | 6381bp |
| 基因突变: | Bacteria codon-optimized nSpCas9 (H840A) & M-MLV RT (D200N, T306K, W313F, T330P, L603W) |
| 启动子: | T7 |
| 标签/融合蛋白: | 6xHis tag (C terminal on insert) |
| 克隆方法: | Gibson Cloning |
| 5’测序引物 | T7 (TAATACGACTCACTATAGGG) |
| 3’测序引物 | T7 Terminal (GCTAGTTATTGCTCAGCGG) |
|
文献引用方法 |
|
| 材料和方法部分: | pET-PE2-His (IK1822) was a gift from Keith Joung (Addgene plasmid # 170103 ; http://n2t.net/addgene:170103 ; RRID:Addgene_170103) |
| 参考文献部分: | CRISPR prime editing with ribonucleoprotein complexes in zebrafish and primary human cells. Petri K, Zhang W, Ma J, Schmidts A, Lee H, Horng JE, Kim DY, Kurt IC, Clement K, Hsu JY, Pinello L, Maus MV, Joung JK, Yeh JJ. Nat Biotechnol. 2021 Apr 29. pii: 10.1038/s41587-021-00901-y. doi: 10.1038/s41587-021-00901-y. 10.1038/s41587-021-00901-y PubMed 33927418 |
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 pET-PE2-His (IK1822) 11619 bp ds-DNA circular SYN 13-MAY-2021
DEFINITION T7 promoter bacterial expression plasmid for PE2
(nSpCas9(H840A)-MMLV-RT) with C-terminal 6xHis-tag and bipartite
NLS.
ACCESSION .
VERSION .
KEYWORDS pET-PE2-His (IK1822)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 11619)
TITLE CRISPR prime editing with ribonucleoprotein complexes in zebrafish
and primary human cells
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 11619)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..11619
/organism="synthetic DNA construct"
/mol_type="other DNA"
rep_origin 57..512
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(144..163)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 354..375
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(604..1419)
/codon_start=1
/gene="aph(3')-Ia"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin in bacteria or G418
(Geneticin(R)) in eukaryotes"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
primer_bind 1327..1346
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
primer_bind 1411..1430
/label=pENTR-R
/note="pENTR vectors, reverse primer"
rep_origin 1541..2129
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 2030..2049
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 2283..2300
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 2315..2454
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(2380..2399)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 2540..2562
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(2556..2747)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(3556..4638)
/codon_start=1
/gene="lacI"
/product="lac repressor"
/label=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)."
/translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
primer_bind 4600..4619
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(4639..4716)
/gene="lacI"
/label=lacI promoter
primer_bind 4922..4941
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 5025..5044
/label=T7
/note="T7 promoter, forward primer"
promoter 5025..5043
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 5044..5068
/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)."
RBS 5083..5105
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 5148..5168
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5169..9269
/codon_start=1
/product="nickase mutant of the Cas9 endonuclease from the
Streptococcus pyogenes Type II CRISPR/Cas system"
/label=Cas9(H840A)
/note="generates RNA-guided single strand nicks in DNA due
to the H840A mutation in the HNH catalytic domain"
/translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 11442..11462
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 11472..11489
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 11508..11525
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(11578..11596)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 11592..20
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
ORIGIN
1 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg aatgggacgc
61 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
121 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
181 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
241 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
301 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
361 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
421 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
481 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
541 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgaattaa
601 ttcttagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc
661 aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt
721 ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca
781 acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac
841 gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg
901 ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga
961 ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat
1021 cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg
1081 atattcttct aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc
1141 atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca
1201 gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag
1261 aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc
1321 gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg
1381 cggcctagag caagacgttt cccgttgaat atggctcata acaccccttg tattactgtt
1441 tatgtaagca gacagtttta ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc
1501 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
1561 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
1621 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
1681 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
1741 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
1801 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
1861 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
1921 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
1981 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
2041 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
2101 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
2161 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
2221 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
2281 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
2341 atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct ctgatgccgc
2401 atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
2461 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
2521 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
2581 aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg
2641 tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt
2701 ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga tgcctccgtg
2761 taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag gatgctcacg
2821 atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg taaacaactg
2881 gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt
2941 aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca gatccggaac
3001 ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg gaaaccgaag
3061 accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc
3121 tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc
3181 ctcaacgaca ggagcacgat catgcgcacc cgtggggccg ccatgccggc gataatggcc
3241 tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc
3301 aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc
3361 tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag
3421 acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg
3481 ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca
3541 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
3601 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt
3661 tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag
3721 ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt
3781 taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatatc
3841 cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc
3901 gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg
3961 aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg
4021 agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc
4081 cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt
4141 accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa
4201 taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg
4261 atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca
4321 ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc
4381 gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc
4441 aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta
4501 attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc
4561 ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta
4621 taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc
4681 cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc tctcccttat
4741 gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg
4801 caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc cccggccacg gggcctgcca
4861 ccatacccac gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga tcttccccat
4921 cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca
4981 cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg aaattaatac gactcactat
5041 aggggaattg tgagcggata acaattcccc tctagaaata attttgttta actttaagaa
5101 ggagatatac catgaaacgg acagccgacg gaagcgagtt cgagtcacca aagaagaagc
5161 ggaaagtcga taagaaatac tcaataggct tagatatcgg cacaaatagc gtcggatggg
5221 cggtgatcac tgatgaatat aaggttccgt ctaaaaagtt caaggttctg ggaaatacag
5281 accgccacag tatcaaaaaa aatcttatag gggctctttt atttgacagt ggagagacag
5341 cggaagcgac tcgtctcaaa cggacagctc gtagaaggta tacacgtcgg aagaatcgta
5401 tttgttatct acaggagatt ttttcaaatg agatggcgaa agtagatgat agtttctttc
5461 atcgacttga agagtctttt ttggtggaag aagacaagaa gcatgaacgt catcctattt
5521 ttggaaatat agtagatgaa gttgcttatc atgagaaata tccaactatc tatcatctgc
5581 gaaaaaaatt ggtagattct actgataaag cggatttgcg cttaatctat ttggccttag
5641 cgcatatgat taagtttcgt ggtcattttt tgattgaggg agatttaaat cctgataata
5701 gtgatgtgga caaactattt atccagttgg tacaaaccta caatcaatta tttgaagaaa
5761 accctattaa cgcaagtgga gtagatgcta aagcgattct ttctgcacga ttgagtaaat
5821 caagacgatt agaaaatctc attgctcagc tccccggtga gaagaaaaat ggcttatttg
5881 ggaatctcat tgctttgtca ttgggtttga cccctaattt taaatcaaat tttgatttgg
5941 cagaagatgc taaattacag ctttcaaaag atacttacga tgatgattta gataatttat
6001 tggcgcaaat tggagatcaa tatgctgatt tgtttttggc agctaagaat ttatcagatg
6061 ctattttact ttcagatatc ctaagagtaa atactgaaat aactaaggct cccctatcag
6121 cttcaatgat taaacgctac gatgaacatc atcaagactt gactctttta aaagctttag
6181 ttcgacaaca acttccagaa aagtataaag aaatcttttt tgatcaatca aaaaacggat
6241 atgcaggtta tattgatggg ggagctagcc aagaagaatt ttataaattt atcaaaccaa
6301 ttttagaaaa aatggatggt actgaggaat tattggtgaa actaaatcgt gaagatttgc
6361 tgcgcaagca acggaccttt gacaacggct ctattcccca tcaaattcac ttgggtgagc
6421 tgcatgctat tttgagaaga caagaagact tttatccatt tttaaaagac aatcgtgaga
6481 agattgaaaa aatcttgact tttcgaattc cttattatgt tggtccattg gcgcgtggca
6541 atagtcgttt tgcatggatg actcggaagt ctgaagaaac aattacccca tggaattttg
6601 aagaagttgt cgataaaggt gcttcagctc aatcatttat tgaacgcatg acaaactttg
6661 ataaaaatct tccaaatgaa aaagtactac caaaacatag tttgctttat gagtatttta
6721 cggtttataa cgaattgaca aaggtcaaat atgttactga aggaatgcga aaaccagcat
6781 ttctttcagg tgaacagaag aaagccattg ttgatttact cttcaaaaca aatcgaaaag
6841 taaccgttaa gcaattaaaa gaagattatt tcaaaaaaat agaatgtttt gatagtgttg
6901 aaatttcagg agttgaagat agatttaatg cttcattagg tacctaccat gatttgctaa
6961 aaattattaa agataaagat tttttggata atgaagaaaa tgaagatatc ttagaggata
7021 ttgttttaac attgacctta tttgaagata gggagatgat tgaggaaaga cttaaaacat
7081 atgctcacct ctttgatgat aaggtgatga aacagcttaa acgtcgccgt tatactggtt
7141 ggggacgttt gtctcgaaaa ttgattaatg gtattaggga taagcaatct ggcaaaacaa
7201 tattagattt tttgaaatca gatggttttg ccaatcgcaa ttttatgcag ctgatccatg
7261 atgatagttt gacatttaaa gaagacattc aaaaagcaca agtgtctgga caaggcgata
7321 gtttacatga acatattgca aatttagctg gtagccctgc tattaaaaaa ggtattttac
7381 agactgtaaa agttgttgat gaattggtca aagtaatggg gcggcataag ccagaaaata
7441 tcgttattga aatggcacgt gaaaatcaga caactcaaaa gggccagaaa aattcgcgag
7501 agcgtatgaa acgaatcgaa gaaggtatca aagaattagg aagtcagatt cttaaagagc
7561 atcctgttga aaatactcaa ttgcaaaatg aaaagctcta tctctattat ctccaaaatg
7621 gaagagacat gtatgtggac caagaattag atattaatcg tttaagtgat tatgatgtcg
7681 atgcaattgt tccacaaagt ttccttaaag acgattcaat agacaataag gtcttaacgc
7741 gttctgataa aaatcgtggt aaatcggata acgttccaag tgaagaagta gtcaaaaaga
7801 tgaaaaacta ttggagacaa cttctaaacg ccaagttaat cactcaacgt aagtttgata
7861 atttaacgaa agctgaacgt ggaggtttga gtgaacttga taaagctggt tttatcaaac
7921 gccaattggt tgaaactcgc caaatcacta agcatgtggc acaaattttg gatagtcgca
7981 tgaatactaa atacgatgaa aatgataaac ttattcgaga ggttaaagtg attaccttaa
8041 aatctaaatt agtttctgac ttccgaaaag atttccaatt ctataaagta cgtgagatta
8101 acaattacca tcatgcccat gatgcgtatc taaatgccgt cgttggaact gctttgatta
8161 agaaatatcc aaaacttgaa tcggagtttg tctatggtga ttataaagtt tatgatgttc
8221 gtaaaatgat tgctaagtct gagcaagaaa taggcaaagc aaccgcaaaa tatttctttt
8281 actctaatat catgaacttc ttcaaaacag aaattacact tgcaaatgga gagattcgca
8341 aacgccctct aatcgaaact aatggggaaa ctggagaaat tgtctgggat aaagggcgag
8401 attttgccac agtgcgcaaa gtattgtcca tgccccaagt caatattgtc aagaaaacag
8461 aagtacagac aggcggattc tccaaggagt caattttacc aaaaagaaat tcggacaagc
8521 ttattgctcg taaaaaagac tgggatccaa aaaaatatgg tggttttgat agtccaacgg
8581 tagcttattc agtcctagtg gttgctaagg tggaaaaagg gaaatcgaag aagttaaaat
8641 ccgttaaaga gttactaggg atcacaatta tggaaagaag ttcctttgaa aaaaatccga
8701 ttgacttttt agaagctaaa ggatataagg aagttaaaaa agacttaatc attaaactac
8761 ctaaatatag tctttttgag ttagaaaacg gtcgtaaacg gatgctggct agtgccggag
8821 aattacaaaa aggaaatgag ctggctctgc caagcaaata tgtgaatttt ttatatttag
8881 ctagtcatta tgaaaagttg aagggtagtc cagaagataa cgaacaaaaa caattgtttg
8941 tggagcagca taagcattat ttagatgaga ttattgagca aatcagtgaa ttttctaagc
9001 gtgttatttt agcagatgcc aatttagata aagttcttag tgcatataac aaacatagag
9061 acaaaccaat acgtgaacaa gcagaaaata ttattcattt atttacgttg acgaatcttg
9121 gagctccagc cgcattcaag tattttgaca caacgataga tcgcaaacga tacacttcta
9181 ccaaggaggt gctagacgcg acactgattc accaatccat cacgggatta tatgaaactc
9241 ggatagattt gtcacagctt gggggtgact ctggaggatc tagcggagga tcctctggca
9301 gcgagacacc aggaacaagc gagtcagcaa caccagagag cagtggcggc agcagcggcg
9361 gcagcagcac cctaaatata gaagatgagt atcggctaca tgagacctca aaagagccag
9421 atgtttctct agggtccaca tggctgtctg attttcctca ggcctgggcg gaaaccgggg
9481 gcatgggact ggcagttcgc caagctcctc tgatcatacc tctgaaagca acctctaccc
9541 ccgtgtccat aaaacaatac cccatgtcac aagaagccag actggggatc aagccccaca
9601 tacagagact gttggaccag ggaatactgg taccctgcca gtccccctgg aacacgcccc
9661 tgctacccgt taagaaacca gggactaatg attataggcc tgtccaggat ctgagagaag
9721 tcaacaagcg ggtggaagac atccacccca ccgtgcccaa cccttacaac ctcttgagcg
9781 ggctcccacc gtcccaccag tggtacactg tgcttgattt aaaggatgcc tttttctgcc
9841 tgagactcca ccccaccagt cagcctctct tcgcctttga gtggagagat ccagagatgg
9901 gaatctcagg acaattgacc tggaccagac tcccacaggg tttcaaaaac agtcccaccc
9961 tgtttaatga ggcactgcac agagacctag cagacttccg gatccagcac ccagacttga
10021 tcctgctaca gtacgtggat gacttactgc tggccgccac ttctgagcta gactgccaac
10081 aaggtactcg ggccctgtta caaaccctag ggaacctcgg gtatcgggcc tcggccaaga
10141 aagcccaaat ttgccagaaa caggtcaagt atctggggta tcttctaaaa gagggtcaga
10201 gatggctgac tgaggccaga aaagagactg tgatggggca gcctactccg aagacccctc
10261 gacaactaag ggagttccta gggaaggcag gcttctgtcg cctcttcatc cctgggtttg
10321 cagaaatggc agcccccctg taccctctca ccaaaccggg gactctgttt aattggggcc
10381 cagaccaaca aaaggcctat caagaaatca agcaagctct tctaactgcc ccagccctgg
10441 ggttgccaga tttgactaag ccctttgaac tctttgtcga cgagaagcag ggctacgcca
10501 aaggtgtcct aacgcaaaaa ctgggacctt ggcgtcggcc ggtggcctac ctgtccaaaa
10561 agctagaccc agtagcagct gggtggcccc cttgcctacg gatggtagca gccattgccg
10621 tactgacaaa ggatgcaggc aagctaacca tgggacagcc actagtcatt ctggcccccc
10681 atgcagtaga ggcactagtc aaacaacccc ccgaccgctg gctttccaac gcccggatga
10741 ctcactatca ggccttgctt ttggacacgg accgggtcca gttcggaccg gtggtagccc
10801 tgaacccggc tacgctgctc ccactgcctg aggaagggct gcaacacaac tgccttgata
10861 tcctggccga agcccacgga acccgacccg acctaacgga ccagccgctc ccagacgccg
10921 accacacctg gtacacggat ggaagcagtc tcttacaaga gggacagcgt aaggcgggag
10981 ctgcggtgac caccgagacc gaggtaatct gggctaaagc cctgccagcc gggacatccg
11041 ctcagcgggc tgaactgata gcactcaccc aggccctaaa gatggcagaa ggtaagaagc
11101 taaatgttta tactgatagc cgttatgctt ttgctactgc ccatatccat ggagaaatat
11161 acagaaggcg tgggtggctc acatcagaag gcaaagagat caaaaataaa gacgagatct
11221 tggccctact aaaagccctc tttctgccca aaagacttag cataatccat tgtccaggac
11281 atcaaaaggg acacagcgcc gaggctagag gcaaccggat ggctgaccaa gcggcccgaa
11341 aggcagccat cacagagact ccagacacct ctaccctcct catagaaaat tcatcaccct
11401 ctggcggctc aaaaagaacc gccgacggca gcgaattcga gcccaagaag aagaggaaag
11461 tcggcagcag ccatcatcat catcatcact aagcggccgc actcgagcac caccaccacc
11521 accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg
11581 ctgagcaata actagcataa ccccttgggg cctctaaac
//
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