| 质粒名称: | pRDA_085 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 158583 |
| 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: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | SpCas9 |
| 物种来源: | Synthetic |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 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_085 was a gift from John Doench & David Root (Addgene plasmid # 158583 ; http://n2t.net/addgene:158583 ; RRID:Addgene_158583) |
买家导航
pRDA_085载体 pRDA085质粒基本信息
pRDA_085质粒图谱 pRDA085载体图谱和pRDA_085载体序列 pRDA085质粒序列多克隆位点信息
pRDA_085质粒 pRDA085载体简介
pRDA_085质粒序列 pRDA085载体序列
LOCUS pRDA_085 12797 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Lentiviral expression of SpCas9; P2A site provides mKate2.
ACCESSION .
VERSION .
KEYWORDS pRDA_085
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 12797)
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 12797)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..12797
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 87..1265
/label=EF-1-alpha promoter
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
intron 318..1256
/label=EF-1-alpha intron A
/note="intron upstream of the start codon of human
EF-1-alpha"
primer_bind 1213..1233
/label=EF1a-F
/note="Human elongation factor-1a promoter, forward primer"
CDS 1282..1302
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 1309..5409
/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 5416..5463
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS 5476..5529
/codon_start=1
/product="2A peptide from Thosea asigna virus capsid
protein"
/label=T2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="EGRGSLLTCGDVEENPGP"
CDS 5536..5931
/codon_start=1
/gene="Aspergillus terreus BSD"
/product="blasticidin S deaminase"
/label=BSD
/note="confers resistance to blasticidin"
/translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
KAIVKDSDGQPTAVGIRELLPSGYVWEG"
CDS 5932..5988
/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 5995..6693
/codon_start=1
/product="monomeric far-red fluorescent protein (Shcherbo
et al., 2009)"
/label=mKate2
/note="mammalian codon-optimized"
/translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
CDLPSKLGHR"
misc_feature 6712..7300
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(6765..6785)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(7183..7194)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
LTR 7372..7605
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 7683..7804
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(7720..7739)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 7774..7793
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
rep_origin 7844..7979
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 7906..7925
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind complement(7999..8018)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(8000..8018)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(8028..8045)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(8028..8044)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(8037..8059)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 8186..8641
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(8273..8292)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 8483..8504
/label=F1ori-F
/note="F1 origin, forward primer"
promoter 8667..8771
/gene="bla"
/label=AmpR promoter
CDS 8772..9632
/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(8990..9009)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 9803..10391
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 10292..10311
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 10545..10562
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 10679..10700
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 10715..10745
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 10753..10769
/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 10758..10780
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 10777..10793
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 10777..10793
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind 10812..10832
/label=T3
/note="T3 promoter, forward primer"
promoter 10814..10832
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
promoter 10860..11086
/label=RSV promoter
/note="Rous sarcoma virus enhancer/promoter"
LTR 11087..11267
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 11314..11439
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 11932..12165
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 12350..12394
/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 12628..12745
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
ORIGIN
1 cccgggtgca aagatggata aagttttaaa cagagaggaa tctttgcagc taatggacct
61 tctaggtctt gaaaggagtg ggaattggct ccggtgcccg tcagtgggca gagcgcacat
121 cgcccacagt ccccgagaag ttggggggag gggtcggcaa ttgaaccggt gcctagagaa
181 ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg
241 gtgggggaga accgtatata agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt
301 ttgccgccag aacacaggta agtgccgtgt gtggttcccg cgggcctggc ctctttacgg
361 gttatggccc ttgcgtgcct tgaattactt ccacctggct gcagtacgtg attcttgatc
421 ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc ttgcgcttaa ggagcccctt
481 cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg ccgccgcgtg cgaatctggt
541 ggcaccttcg cgcctgtctc gctgctttcg ataagtctct agccatttaa aatttttgat
601 gacctgctgc gacgcttttt ttctggcaag atagtcttgt aaatgcgggc caagatctgc
661 acactggtat ttcggttttt ggggccgcgg gcggcgacgg ggcccgtgcg tcccagcgca
721 catgttcggc gaggcggggc ctgcgagcgc ggccaccgag aatcggacgg gggtagtctc
781 aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc gtgtatcgcc ccgccctggg
841 cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga aagatggccg cttcccggcc
901 ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg agagcgggcg ggtgagtcac
961 ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc ttcatgtgac tccacggagt
1021 accgggcgcc gtccaggcac ctcgattagt tctcgagctt ttggagtacg tcgtctttag
1081 gttgggggga ggggttttat gcgatggagt ttccccacac tgagtgggtg gagactgaag
1141 ttaggccagc ttggcacttg atgtaattct ccttggaatt tgcccttttt gagtttggat
1201 cttggttcat tctcaagcct cagacagtgg ttcaaagttt ttttcttcca tttcaggtgt
1261 cgtgagggat ccaccatggc accaaagaag aagcggaagg tcgcagccga caagaagtac
1321 agcatcggcc tggacatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac
1381 aaggtgccca gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag
1441 aacctgatcg gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag
1501 agaaccgcca gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc
1561 ttcagcaacg agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc
1621 ctggtggaag aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag
1681 gtggcctacc acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc
1741 accgacaagg ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg
1801 ggccacttcc tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc
1861 atccagctgg tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc
1921 gtagacgcca aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg
1981 atcgcccagc tgcccggcga gaagaagaat ggcctgttcg gaaacctgat tgccctgagc
2041 ctgggcctga cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag
2101 ctgagcaagg acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag
2161 tacgccgacc tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc
2221 ctgagagtga acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac
2281 gacgagcacc accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag
2341 aagtacaaag agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc
2401 ggagccagcc aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc
2461 accgaggaac tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcgtacgttc
2521 gacaacggca gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg
2581 caggaagatt tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc
2641 ttccgcatcc cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg
2701 accagaaaga gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc
2761 gctagcgccc agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag
2821 aaggtgctgc ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc
2881 aaagtgaaat acgtgaccga gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa
2941 aaggccatcg tggacctgct gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa
3001 gaggactact tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg cgtggaagat
3061 cggttcaacg cctccctggg cacataccac gatctgctga aaattatcaa ggacaaggac
3121 ttcctggaca atgaggaaaa cgaggacatt ctggaagata tcgtgctgac cctgacactg
3181 tttgaggaca gagagatgat cgaggaacgg ctgaaaacct atgcccacct gttcgacgac
3241 aaagtgatga agcagctgaa gcggcggaga tacaccggct ggggcaggct gagccggaag
3301 ctgatcaacg gcatccggga caagcagtcc ggcaagacaa tcctggattt cctgaagtcc
3361 gacggcttcg ccaacagaaa cttcatgcag ctgatccacg acgacagcct gacctttaaa
3421 gaggacatcc agaaagccca ggtgtccggc cagggcgata gcctgcacga gcacattgcc
3481 aatctggccg gcagccccgc cattaagaag ggcatcctgc agacagtgaa ggtggtggac
3541 gagctcgtga aagtgatggg ccggcacaag cccgagaaca tcgtgatcga aatggccaga
3601 gagaaccaga ccacccagaa gggacagaag aacagccgcg agagaatgaa gcggatcgaa
3661 gagggcatca aagagctggg cagccagatc ctgaaagaac accccgtgga aaacacccag
3721 ctgcagaacg agaagctgta cctgtactac ctgcagaatg ggcgggatat gtacgtggac
3781 caggaactgg acatcaaccg gctgtccgac tacgatgtgg accatatcgt gcctcagagc
3841 tttctgaagg acgactccat cgacaacaag gtgctgacca gaagcgacaa gaaccggggc
3901 aagagcgaca acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag
3961 ctgctgaacg ccaagctgat tacccagaga aagttcgaca atctgaccaa ggccgagaga
4021 ggcggcctga gcgaactgga taaggccggc ttcatcaaga gacagctggt ggaaacccgg
4081 cagatcacaa agcacgtggc acagatcctg gactcccgga tgaacactaa gtacgacgag
4141 aatgacaagc tgatccggga agtgaaagtg atcaccctga agtccaagct ggtgtccgat
4201 ttccggaagg atttccagtt ttacaaagtg cgcgagatca acaactacca ccacgcccac
4261 gacgcctacc tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc taagctggaa
4321 agcgagttcg tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc
4381 gagcaggaaa tcggcaaggc taccgccaag tacttcttct acagcaacat catgaacttt
4441 ttcaagaccg agattaccct ggccaacggc gagatccgga agcggcctct gatcgagaca
4501 aacggcgaaa ccggggagat cgtgtgggat aagggccggg attttgccac cgtgcggaaa
4561 gtgctgagca tgccccaagt gaatatcgtg aaaaagaccg aggtgcagac aggcggcttc
4621 agcaaagagt ctatcctgcc caagaggaac agcgataagc tgatcgccag aaagaaggac
4681 tgggacccta agaagtacgg cggcttcgac agccccaccg tggcctattc tgtgctggtg
4741 gtggccaaag tggaaaaggg caagtccaag aaactgaaga gtgtgaaaga gctgctgggg
4801 atcaccatca tggaaagaag cagcttcgag aagaatccca tcgactttct ggaagccaag
4861 ggctacaaag aagtgaaaaa ggacctgatc atcaagctgc ctaagtactc cctgttcgag
4921 ctggaaaacg gccggaagag aatgctggcc tctgccggcg aactgcagaa gggaaacgaa
4981 ctggccctgc cctccaaata tgtgaacttc ctgtacctgg ccagccacta tgagaagctg
5041 aagggctccc ccgaggataa tgagcagaaa cagctgtttg tggaacagca caagcactac
5101 ctggacgaga tcatcgagca gatcagcgag ttctccaaga gagtgatcct ggccgacgct
5161 aatctggaca aagtgctgtc cgcctacaac aagcaccggg ataagcccat cagagagcag
5221 gccgagaata tcatccacct gtttaccctg accaatctgg gagcccctgc cgccttcaag
5281 tactttgaca ccaccatcga ccggaagagg tacaccagca ccaaagaggt gctggacgcc
5341 accctgatcc accagagcat caccggcctg tacgagacac gaatcgattt gtctcagctg
5401 ggaggcgacg ccgcaaaaag gccggcggcc acgaaaaagg ccggccaggc aaaaaagaaa
5461 aaggcaggct caggcgaggg cagaggaagt ctgctaacat gcggtgacgt cgaggagaat
5521 cctggcccaa gcgctatggc caagcctttg tctcaagaag aatccaccct cattgaaaga
5581 gcaacggcta caatcaacag catccccatc tctgaagact acagcgtcgc cagcgcagct
5641 ctctctagcg acggccgcat cttcactggt gtcaatgtat atcattttac tgggggacct
5701 tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg cagctggcaa cctgacttgt
5761 atcgtcgcga tcggaaatga gaacaggggc atcttgagcc cctgcggacg gtgccgacag
5821 gtgcttctcg atctgcatcc tgggatcaaa gccatagtga aggacagtga tggacagccg
5881 acggcagttg ggattcgtga attgctgccc tctggttatg tgtgggaggg cgcaacaaac
5941 ttctctctgc tgaaacaagc cggagatgtc gaagagaatc ctggaccgac tagtatggtg
6001 agcgagctga ttaaggagaa catgcacatg aagctataca tggagggcac cgtgaacaac
6061 caccacttca agtgcacatc cgagggcgaa ggcaagccct acgagggcac ccagaccatg
6121 agaatcaagg cggtcgaggg cggccctctc cccttcgcct tcgacatcct ggctaccagc
6181 ttcatgtacg gcagcaaaac cttcatcaac cacacccagg gcatccccga cttctttaag
6241 cagtccttcc ccgagggctt cacatgggag agagtcacca catacgaaga cgggggcgtg
6301 ctgaccgcta cccaggacac cagcctccag gacggctgcc tcatctacaa cgtcaagatc
6361 agaggggtga acttcccatc caacggccct gtgatgcaga agaaaacact cggctgggag
6421 gcctccaccg agaccctgta ccccgctgac ggcggcctgg aaggcagagc cgacatggcc
6481 ctgaagctcg tgggcggggg ccacctgatc tgcaacttga agaccacata cagatccaag
6541 aaacccgcta agaacctcaa gatgcccggc gtctactatg tggacagaag actggaaaga
6601 atcaaggagg ccgacaaaga gacctacgtc gagcagcacg aggtggctgt ggccagatac
6661 tgcgacctcc ctagcaaact ggggcacaga tgatagacgc gttaagtcga caatcaacct
6721 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
6781 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
6841 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
6901 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
6961 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
7021 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
7081 gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt
7141 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
7201 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
7261 cctcagacga gtcggatctc cctttgggcc gcctccccgc gtcgacttta agaccaatga
7321 cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc
7381 taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag
7441 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
7501 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
7561 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtacg tatagtagtt
7621 catgtcatct tattattcag tatttataac ttgcaaagaa atgaatatca gagagtgaga
7681 ggaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
7741 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
7801 cttatcatgt ctggctctag ctatcccgcc cctaactccg cccatcccgc ccctaactcc
7861 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
7921 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
7981 agggacgtac ccaattcgcc ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt
8041 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
8101 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
8161 ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt
8221 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
8281 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
8341 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
8401 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
8461 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
8521 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
8581 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt
8641 taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
8701 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
8761 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
8821 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
8881 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
8941 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
9001 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
9061 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
9121 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
9181 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
9241 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
9301 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
9361 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
9421 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
9481 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
9541 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
9601 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
9661 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
9721 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
9781 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
9841 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
9901 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
9961 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
10021 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
10081 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
10141 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
10201 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
10261 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
10321 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
10381 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
10441 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
10501 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
10561 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
10621 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
10681 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
10741 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
10801 acgccaagcg cgcaattaac cctcactaaa gggaacaaaa gctggagctg caagcttaat
10861 gtagtcttat gcaatactct tgtagtcttg caacatggta acgatgagtt agcaacatgc
10921 cttacaagga gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg
10981 tgccttatta ggaaggcaac agacgggtct gacatggatt ggacgaacca ctgaattgcc
11041 gcattgcaga gatattgtat ttaagtgcct agctcgatac ataaacgggt ctctctggtt
11101 agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca
11161 ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa
11221 ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagtg gcgcccgaac
11281 agggacttga aagcgaaagg gaaaccagag gagctctctc gacgcaggac tcggcttgct
11341 gaagcgcgca cggcaagagg cgaggggcgg cgactggtga gtacgccaaa aattttgact
11401 agcggaggct agaaggagag agatgggtgc gagagcgtca gtattaagcg ggggagaatt
11461 agatcgcgat gggaaaaaat tcggttaagg ccagggggaa agaaaaaata taaattaaaa
11521 catatagtat gggcaagcag ggagctagaa cgattcgcag ttaatcctgg cctgttagaa
11581 acatcagaag gctgtagaca aatactggga cagctacaac catcccttca gacaggatca
11641 gaagaactta gatcattata taatacagta gcaaccctct attgtgtgca tcaaaggata
11701 gagataaaag acaccaagga agctttagac aagatagagg aagagcaaaa caaaagtaag
11761 accaccgcac agcaagcggc cgctgatctt cagacctgga ggaggagata tgagggacaa
11821 ttggagaagt gaattatata aatataaagt agtaaaaatt gaaccattag gagtagcacc
11881 caccaaggca aagagaagag tggtgcagag agaaaaaaga gcagtgggaa taggagcttt
11941 gttccttggg ttcttgggag cagcaggaag cactatgggc gcagcgtcaa tgacgctgac
12001 ggtacaggcc agacaattat tgtctggtat agtgcagcag cagaacaatt tgctgagggc
12061 tattgaggcg caacagcatc tgttgcaact cacagtctgg ggcatcaagc agctccaggc
12121 aagaatcctg gctgtggaaa gatacctaaa ggatcaacag ctcctgggga tttggggttg
12181 ctctggaaaa ctcatttgca ccactgctgt gccttggaat gctagttgga gtaataaatc
12241 tctggaacag atttggaatc acacgacctg gatggagtgg gacagagaaa ttaacaatta
12301 cacaagctta atacactcct taattgaaga atcgcaaaac cagcaagaaa agaatgaaca
12361 agaattattg gaattagata aatgggcaag tttgtggaat tggtttaaca taacaaattg
12421 gctgtggtat ataaaattat tcataatgat agtaggaggc ttggtaggtt taagaatagt
12481 ttttgctgta ctttctatag tgaatagagt taggcaggga tattcaccat tatcgtttca
12541 gacccacctc ccaaccccga ggggaccctg gtacccatgc atgcatgaat tctagatctt
12601 gagacaaatg gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag
12661 tgcaggggaa agaatagtag acataatagc aacagacata caaactaaag aattacaaaa
12721 acaaattaca aaaattcaaa attttcgggt ttattacagg gacagcagag atccactttg
12781 gcgccggctc gaggggg
//
