| 质粒名称: | pRDA_256 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 158581 |
| 存储实验室: | John Doench, David Root |
|
载体骨架基本信息 |
|
| 载体名称: | pXPR |
| 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: | -- |
| 载体修饰: | -- |
| 载体类型: | 哺乳动物表达，慢病毒 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 30 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | -- |
| 物种来源: | -- |
| 插入大小: | -- |
| 基因突变: | Cas9 D10A |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | -- |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | -- |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pRDA_256 was a gift from John Doench & David Root (Addgene plasmid # 158581 ; http://n2t.net/addgene:158581 ; RRID:Addgene_158581) |
买家导航
pRDA256载体基本信息
pRDA256载体图谱质粒图谱和多克隆位点信息
pRDA256载体简介
pRDA256载体序列
LOCUS pRDA_256 14128 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Lentiviral expression of BE3.9max and customizable gRNA.
ACCESSION .
VERSION .
KEYWORDS pRDA_256
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 14128)
AUTHORS Hanna RE, Hegde M, Fagre CR, DeWeirdt PC, Sangree AK, Szegletes Z,
Griffith A, Feeley MN, Sanson KR, Baidi Y, Koblan LW, Liu DR, Neal
JT, Doench JG
TITLE Massively parallel assessment of human variants with base editor
screens
JOURNAL BioRxiv
REFERENCE 2 (bases 1 to 14128)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..14128
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 108..128
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
LTR 171..351
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 398..523
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1016..1249
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 1434..1478
/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 1776..1893
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 1944..2184
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 1944..1964
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 2115..2134
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 2214..2289
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter 2373..2584
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
CDS 2634..2654
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 2655..3338
/codon_start=1
/product="cytidine deaminase (C to U editing enzyme) from
rat"
/label=APOBEC-1
/note="can use ssDNA as a substrate (Komor et al., 2016)"
/translation="SSETGPVAVDPTLRRRIEPHEFEVFFDPRELRKETCLLYEINWGG
RHSIWRHTSQNTNKHVEVNFIEKFTTERYFCPNTRCSITWFLSWSPCGECSRAITEFLS
RYPHVTLFIYIARLYHHADPRNRQGLRDLISSGVTIQIMTEQESGYCWRNFVNYSPSNE
AHWPRYPHLWVRLYVLELYCIILGLPPCLNILRRKQPQLTFFTIALQSCHYQRLPPHIL
WATGLK"
CDS 3435..7535
/codon_start=1
/product="nickase mutant of the Cas9 endonuclease from the
Streptococcus pyogenes Type II CRISPR/Cas system"
/label=Cas9(D10A)
/note="generates RNA-guided single strand nicks in DNA due
to the D10A mutation in the RuvC catalytic domain"
/translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 7566..7814
/codon_start=1
/product="uracil-DNA glycosylase inhibitor from a Bacillus
subtilis bacteriophage (Mol et al., 1995)"
/label=UGI
/translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
CDS 7857..7877
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 7887..7943
/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 7944..8540
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind 8437..8457
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
misc_feature 8556..9144
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(8609..8629)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(9027..9038)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
LTR 9216..9449
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
primer_bind complement(9475..9492)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 9481..9705
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 9751..10179
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(9838..9857)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 10048..10069
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(10188..10208)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 10193..10522
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 10373..10508
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 10435..10454
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
promoter 10570..10617
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 10636..11010
/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 11140..11261
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(11177..11196)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 11231..11250
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(11310..11326)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(11310..11326)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(11323..11345)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 11334..11350
/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(11358..11388)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 11403..11424
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(11541..11558)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(11712..12300)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(11792..11811)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(12471..13331)
/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 13094..13113
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(13332..13436)
/gene="bla"
/label=AmpR promoter
primer_bind complement(13511..13530)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 13702..14081
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 14083..153
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
ORIGIN
1 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
61 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
121 taggcgtgta cggtgggagg tctatataag cagcgcgttt tgcctgtact gggtctctct
181 ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc
241 ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg
301 gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc
361 gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct
421 tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt
481 gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag
541 aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt
601 aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt
661 agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg
721 atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag
781 gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag
841 taagaccacc gcacagcaag cggccgctga tcttcagacc tggaggagga gatatgaggg
901 acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag
961 cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag
1021 ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc
1081 tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga
1141 gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc
1201 aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg
1261 gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata
1321 aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca
1381 attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg
1441 aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa
1501 attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa
1561 tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt
1621 ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag
1681 gtggagagag agacagagac agatccattc gattagtgaa cggatcggca ctgcgtgcgc
1741 caattctgca gacaaatggc agtattcatc cacaatttta aaagaaaagg ggggattggg
1801 gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca aactaaagaa
1861 ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga cagcagagat
1921 ccagtttggt taattaaggt accgagggcc tatttcccat gattccttca tatttgcata
1981 tacgatacaa ggctgttaga gagataatta gaattaattt gactgtaaac acaaagatat
2041 tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat
2101 tatgttttaa aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg
2161 ctttatatat cttgtggaaa ggacgaaaca ccggagacgc ttttttcgtc tcagtttgag
2221 agctagaaat agcaagttca aataaggcta gtccgttatc aacttgaaaa agtggcaccg
2281 agtcggtgcg ctcacctatt agcggctaag gttttttgaa ttcgctagct aggtcttgaa
2341 aggagtggga attggctccg gtgcccgtca gtgggcagag cgcacatcgc ccacagtccc
2401 cgagaagttg gggggagggg tcggcaattg atccggtgcc tagagaaggt ggcgcggggt
2461 aaactgggaa agtgatgtcg tgtactggct ccgccttttt cccgagggtg ggggagaacc
2521 gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg ccgccagaac
2581 acaggaccgg tgccaccatg aaacggacag ccgacggaag cgagttcgag tcaccaaaga
2641 agaagcggaa agtctcctca gagactgggc ctgtcgccgt cgatccaacc ctgcgccgcc
2701 ggattgaacc tcacgagttt gaagtgttct ttgacccccg ggagctgaga aaggagacat
2761 gcctgctgta cgagatcaac tggggaggca ggcactccat ctggaggcac acctctcaga
2821 acacaaataa gcacgtggag gtgaacttca tcgagaagtt taccacagag cggtacttct
2881 gccccaatac cagatgtagc atcacatggt ttctgagctg gtccccttgc ggagagtgta
2941 gcagggccat caccgagttc ctgtccagat atccacacgt gacactgttt atctacatcg
3001 ccaggctgta tcaccacgca gacccaagga ataggcaggg cctgcgcgat ctgatcagct
3061 ccggcgtgac catccagatc atgacagagc aggagtccgg ctactgctgg cggaacttcg
3121 tgaattattc tcctagcaac gaggcccact ggcctaggta cccacacctg tgggtgcgcc
3181 tgtacgtgct ggagctgtat tgcatcatcc tgggcctgcc cccttgtctg aatatcctgc
3241 ggagaaagca gccccagctg accttcttta caatcgccct gcagtcttgt cactatcaga
3301 ggctgccacc ccacatcctg tgggccacag gcctgaagtc tggaggatct agcggaggat
3361 cctctggcag cgagacacca ggaacaagcg agtcagcaac accagagagc agtggcggca
3421 gcagcggcgg cagcgacaag aagtacagca tcggcctggc catcggcacc aactctgtgg
3481 gctgggccgt gatcaccgac gagtacaagg tgcccagcaa gaaattcaag gtgctgggca
3541 acaccgaccg gcacagcatc aagaagaacc tgatcggagc cctgctgttc gacagcggcg
3601 aaacagccga ggccacccgg ctgaagagaa ccgccagaag aagatacacc agacggaaga
3661 accggatctg ctatctgcaa gagatcttca gcaacgagat ggccaaggtg gacgacagct
3721 tcttccacag actggaagag tccttcctgg tggaagagga taagaagcac gagcggcacc
3781 ccatcttcgg caacatcgtg gacgaggtgg cctaccacga gaagtacccc accatctacc
3841 acctgagaaa gaaactggtg gacagcaccg acaaggccga cctgcggctg atctatctgg
3901 ccctggccca catgatcaag ttccggggcc acttcctgat cgagggcgac ctgaaccccg
3961 acaacagcga cgtggacaag ctgttcatcc agctggtgca gacctacaac cagctgttcg
4021 aggaaaaccc catcaacgcc agcggcgtgg acgccaaggc catcctgtct gccagactga
4081 gcaagagcag acggctggaa aatctgatcg cccagctgcc cggcgagaag aagaatggcc
4141 tgttcggaaa cctgattgcc ctgagcctgg gcctgacccc caacttcaag agcaacttcg
4201 acctggccga ggatgccaaa ctgcagctga gcaaggacac ctacgacgac gacctggaca
4261 acctgctggc ccagatcggc gaccagtacg ccgacctgtt tctggccgcc aagaacctgt
4321 ccgacgccat cctgctgagc gacatcctga gagtgaacac cgagatcacc aaggcccccc
4381 tgagcgcctc tatgatcaag agatacgacg agcaccacca ggacctgacc ctgctgaaag
4441 ctctcgtgcg gcagcagctg cctgagaagt acaaagagat tttcttcgac cagagcaaga
4501 acggctacgc cggctacatt gacggcggag ccagccagga agagttctac aagttcatca
4561 agcccatcct ggaaaagatg gacggcaccg aggaactgct cgtgaagctg aacagagagg
4621 acctgctgcg gaagcagcgg accttcgaca acggcagcat cccccaccag atccacctgg
4681 gagagctgca cgccattctg cggcggcagg aagattttta cccattcctg aaggacaacc
4741 gggaaaagat cgagaagatc ctgaccttcc gcatccccta ctacgtgggc cctctggcca
4801 ggggaaacag cagattcgcc tggatgacca gaaagagcga ggaaaccatc accccctgga
4861 acttcgagga agtggtggac aagggcgctt ccgcccagag cttcatcgag cggatgacca
4921 acttcgataa gaacctgccc aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt
4981 acttcaccgt gtataacgag ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc
5041 ccgccttcct gagcggcgag cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc
5101 ggaaagtgac cgtgaagcag ctgaaagagg actacttcaa gaaaatcgag tgcttcgact
5161 ccgtggaaat ctccggcgtg gaagatcggt tcaacgcctc cctgggcaca taccacgatc
5221 tgctgaaaat tatcaaggac aaggacttcc tggacaatga ggaaaacgag gacattctgg
5281 aagatatcgt gctgaccctg acactgtttg aggacagaga gatgatcgag gaacggctga
5341 aaacctatgc ccacctgttc gacgacaaag tgatgaagca gctgaagcgg cggagataca
5401 ccggctgggg caggctgagc cggaagctga tcaacggcat ccgggacaag cagtccggca
5461 agacaatcct ggatttcctg aagtccgacg gcttcgccaa cagaaacttc atgcagctga
5521 tccacgacga cagcctgacc tttaaagagg acatccagaa agcccaggtg tccggccagg
5581 gcgatagcct gcacgagcac attgccaatc tggccggcag ccccgccatt aagaagggca
5641 tcctgcagac agtgaaggtg gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg
5701 agaacatcgt gatcgaaatg gccagagaga accagaccac ccagaaggga cagaagaaca
5761 gccgcgagag aatgaagcgg atcgaagagg gcatcaaaga gctgggcagc cagatcctga
5821 aagaacaccc cgtggaaaac acccagctgc agaacgagaa gctgtacctg tactacctgc
5881 agaatgggcg ggatatgtac gtggaccagg aactggacat caaccggctg tccgactacg
5941 atgtggacca tatcgtgcct cagagctttc tgaaggacga ctccatcgac aacaaggtgc
6001 tgaccagaag cgacaagaac cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga
6061 agaagatgaa gaactactgg cggcagctgc tgaacgccaa gctgattacc cagagaaagt
6121 tcgacaatct gaccaaggcc gagagaggcg gcctgagcga actggataag gccggcttca
6181 tcaagagaca gctggtggaa acccggcaga tcacaaagca cgtggcacag atcctggact
6241 cccggatgaa cactaagtac gacgagaatg acaagctgat ccgggaagtg aaagtgatca
6301 ccctgaagtc caagctggtg tccgatttcc ggaaggattt ccagttttac aaagtgcgcg
6361 agatcaacaa ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc
6421 tgatcaaaaa gtaccctaag ctggaaagcg agttcgtgta cggcgactac aaggtgtacg
6481 acgtgcggaa gatgatcgcc aagagcgagc aggaaatcgg caaggctacc gccaagtact
6541 tcttctacag caacatcatg aactttttca agaccgagat taccctggcc aacggcgaga
6601 tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg
6661 gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa
6721 agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag aggaacagcg
6781 ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc ttcgacagcc
6841 ccaccgtggc ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac
6901 tgaagagtgt gaaagagctg ctggggatca ccatcatgga aagaagcagc ttcgagaaga
6961 atcccatcga ctttctggaa gccaagggct acaaagaagt gaaaaaggac ctgatcatca
7021 agctgcctaa gtactccctg ttcgagctgg aaaacggccg gaagagaatg ctggcctctg
7081 ccggcgaact gcagaaggga aacgaactgg ccctgccctc caaatatgtg aacttcctgt
7141 acctggccag ccactatgag aagctgaagg gctcccccga ggataatgag cagaaacagc
7201 tgtttgtgga acagcacaag cactacctgg acgagatcat cgagcagatc agcgagttct
7261 ccaagagagt gatcctggcc gacgctaatc tggacaaagt gctgtccgcc tacaacaagc
7321 accgggataa gcccatcaga gagcaggccg agaatatcat ccacctgttt accctgacca
7381 atctgggagc ccctgccgcc ttcaagtact ttgacaccac catcgaccgg aagaggtaca
7441 ccagcaccaa agaggtgctg gacgccaccc tgatccacca gagcatcacc ggcctgtacg
7501 agacacggat cgacctgtct cagctgggag gtgacagcgg cgggagcggc gggagcgggg
7561 ggagcactaa tctgagcgac atcattgaga aggagactgg gaaacagctg gtcattcagg
7621 agtccatcct gatgctgcct gaggaggtgg aggaagtgat cggcaacaag ccagagtctg
7681 acatcctggt gcacaccgcc tacgacgagt ccacagatga gaatgtgatg ctgctgacct
7741 ctgacgcccc cgagtataag ccttgggccc tggtcatcca ggattctaac ggcgagaata
7801 agatcaagat gctgagcgga ggctccaaaa gaaccgccga cggcagcgag ttcgagccca
7861 agaagaagag gaaagtcagc gctggcgcaa caaacttctc tctgctgaaa caagccggag
7921 atgtcgaaga gaatcctgga ccgaccgagt acaagcccac ggtgcgcctc gccacccgcg
7981 acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
8041 gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
8101 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
8161 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
8221 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
8281 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggagtc tcgcccgacc
8341 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
8401 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
8461 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
8521 cccgcaagcc cggtgcctga acgcgttaag tcgacaatca acctctggat tacaaaattt
8581 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
8641 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
8701 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
8761 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
8821 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
8881 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
8941 tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc
9001 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
9061 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
9121 tctccctttg ggccgcctcc ccgcgtcgac tttaagacca atgacttaca aggcagctgt
9181 agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg
9241 aagacaagat ctgctttttg cttgtactgg gtctctctgg ttagaccaga tctgagcctg
9301 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
9361 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
9421 cttttagtca gtgtggaaaa tctctagcag ggcccgttta aacccgctga tcagcctcga
9481 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
9541 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
9601 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
9661 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
9721 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
9781 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
9841 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
9901 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
9961 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
10021 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
10081 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
10141 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
10201 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
10261 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
10321 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
10381 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
10441 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
10501 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
10561 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
10621 gtgaggaact aaaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac
10681 gtcgccggag cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag
10741 gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac
10801 caggtggtgc cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac
10861 gccgagtggt cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc
10921 gagatcggcg agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc
10981 gtgcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga ttccaccgcc
11041 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
11101 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
11161 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
11221 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
11281 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
11341 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
11401 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
11461 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
11521 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
11581 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
11641 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
11701 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
11761 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
11821 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
11881 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
11941 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
12001 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
12061 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
12121 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
12181 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
12241 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
12301 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
12361 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
12421 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
12481 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
12541 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
12601 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
12661 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
12721 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
12781 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
12841 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
12901 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
12961 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
13021 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
13081 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
13141 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
13201 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
13261 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
13321 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
13381 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
13441 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
13501 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
13561 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
13621 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
13681 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
13741 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
13801 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
13861 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
13921 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
13981 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
14041 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
14101 acatcaatgg gcgtggatag cggtttga
//
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