| 质粒名称: | Lenti-(BB)-hPGK-KRAB-dCas9-P2A-BlastR |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 118155 |
| 存储实验室: | Jorge Ferrer |
|
载体骨架基本信息 |
|
| 载体名称: | lentiCRISPRv2 |
| 载体出品公司: | Feng Zhang (Addgene #52961) |
| 载体抗性: | 氨苄青霉素 |
| 载体类型: | 哺乳动物表达，慢病毒，CRISPR |
| 筛选标记: | 杀螨素 |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | KRAB-dCas9-P2A-BlastR |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 4908 bp |
| 启动子 | hPGK and U6 |
| 标签/融合蛋白: | HA（插入件上的C端子） |
| 克隆方法: | 限制性内切酶 |
| 5' 测序引物 | CTTGCTGGGCACCTTGTACT |
|
文献引用方法 |
|
| 材料和方法部分: | Lenti-(BB)-hPGK-KRAB-dCas9-P2A-BlastR was a gift from Jorge Ferrer (Addgene plasmid # 118155 ; http://n2t.net/addgene:118155 ; RRID:Addgene_118155) |
| 参考文献部分: | Human pancreatic islet three-dimensional chromatin architecture provides insights into the genetics of type 2 diabetes. Miguel-Escalada I, Bonas-Guarch S, Cebola I, Ponsa-Cobas J, Mendieta-Esteban J, Atla G, Javierre BM, Rolando DMY, Farabella I, Morgan CC, Garcia-Hurtado J, Beucher A, Moran I, Pasquali L, Ramos-Rodriguez M, Appel EVR, Linneberg A, Gjesing AP, Witte DR, Pedersen O, Grarup N, Ravassard P, Torrents D, Mercader JM, Piemonti L, Berney T, de Koning EJP, Kerr-Conte J, Pattou F, Fedko IO, Groop L, Prokopenko I, Hansen T, Marti-Renom MA, Fraser P, Ferrer J. Nat Genet. 2019 Jun 28. pii: 10.1038/s41588-019-0457-0. doi: 10.1038/s41588-019-0457-0. 10.1038/s41588-019-0457-0 PubMed 31253982 |
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
-hPGK-KRAB-dCas9-P2A-BlastR质粒图谱.png)
载体简介
载体序列
LOCUS Lenti-(BB)-hPGK-KRAB-dCas9-P2A-BlastR 15243 bp ds-DNA circular SYN 06-MAY-2019
DEFINITION Catalytically inactive Cas9 from S. pyogenes with P2A-BlastR under
the hPGK promoter, and cloning backbone for sgRNA. Contains BsmBI
sites for insertion of spacer sequences..
ACCESSION .
VERSION .
KEYWORDS Lenti-(BB)-hPGK-KRAB-dCas9-P2A-BlastR
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 15243)
AUTHORS Miguel-Escalada I, Bonas-Guarch S , Cebola I, Ponsa-Cobas J,
Mendieta-Esteban J, Rolando DMY, Javierre BM, Atla G, Farabella I,
Morgan CC, Garcia-Hurtado J, Beucher A, Moran I, Pasquali L, Ramos
M, Appel EVR, Linneberg A, Gjesing AP, Witte DR, Pedersen O, Garup
N, Ravassard P, Torrents D, Mercader JM, Piemonti L, Berney T, de
Koning EJP, Kerr-Conte J, Pattou F, Fedko IO, Prokopenko I, Hansen
T, Marti-Renom MA, Fraser P, Ferrer J
TITLE Human pancreatic islet 3D chromatin architecture provides insights
into the genetics of type 2 diabetes
JOURNAL bioRxiv 400291
REFERENCE 2 (bases 1 to 15243)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 6, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..15243
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 14..524
/label=hPGK promoter
/note="human phosphoglycerate kinase 1 promoter"
primer_bind 443..461
/label=hPGK-F
/note="Human PGK promoter, forward primer"
regulatory 537..546
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 573..758
/codon_start=1
/gene="ZNF10"
/gene_synonym="KOX1"
/product="Kruppel-associated box (KRAB) transcriptional
repression domain from the human zinc finger protein ZNF10
(Margolin et al., 1994)"
/label=KRAB
/translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
QLTKPDVILRLEKGEEP"
CDS 780..2498
/codon_start=1
/product="N-terminal portion of Streptococcus pyogenes Cas9
(Zetsche et al., 2015)"
/label=Cas9(N)
/translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIE"
CDS 4887..4913
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
CDS 4932..4952
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 4959..4979
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5025..5081
/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 5082..5480
/codon_start=1
/gene="Aspergillus terreus BSD"
/product="blasticidin S deaminase"
/label=BSD
/note="confers resistance to blasticidin"
/translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
KAIVKDSDGQPTAVGIRELLPSGYVWEG"
misc_feature 5505..6093
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(5558..5578)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(5976..5987)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
LTR 6165..6398
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
primer_bind complement(6424..6441)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 6430..6654
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 6700..7128
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(6787..6806)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 6997..7018
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(7137..7157)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 7142..7471
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 7322..7457
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 7384..7403
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
promoter 7519..7566
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 7585..7959
/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 8089..8210
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(8126..8145)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 8180..8199
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(8259..8275)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(8259..8275)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(8272..8294)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 8283..8299
/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(8307..8337)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 8352..8373
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(8490..8507)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(8661..9249)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(8741..8760)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(9420..10280)
/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 10043..10062
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(10281..10385)
/gene="bla"
/label=AmpR promoter
primer_bind complement(10460..10479)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 10651..11030
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 11032..11230
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 11185..11205
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
LTR 11248..11428
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 11475..11600
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 12093..12326
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
misc_feature 12853..12970
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 13021..13261
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 13021..13041
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 13192..13211
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 15151..15226
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
ORIGIN
1 ctaggtcttg aaaggggttg gggttgcgcc ttttccaagg cagccctggg tttgcgcagg
61 gacgcggctg ctctgggcgt ggttccggga aacgcagcgg cgccgaccct gggtctcgca
121 cattcttcac gtccgttcgc agcgtcaccc ggatcttcgc cgctaccctt gtgggccccc
181 cggcgacgct tcctgctccg cccctaagtc gggaaggttc cttgcggttc gcggcgtgcc
241 ggacgtgaca aacggaagcc gcacgtctca ctagtaccct cgcagacgga cagcgccagg
301 gagcaatggc agcgcgccga ccgcgatggg ctgtggccaa tagcggctgc tcagcagggc
361 gcgccgagag cagcggccgg gaaggggcgg tgcgggaggc ggggtgtggg gcggtagtgt
421 gggccctgtt cctgcccgcg cggtgttccg cattctgcaa gcctccggag cgcacgtcgg
481 cagtcggctc cctcgttgac cgaatcaccg acctctctcc ccaggaccgg tgatccgcca
541 ccatggatgc taagtcacta actgcctggt cccggacact ggtgaccttc aaggatgtat
601 ttgtggactt caccagggag gagtggaagc tgctggacac tgctcagcag atcgtgtaca
661 gaaatgtgat gctggagaac tataagaacc tggtttcctt gggttatcag cttactaagc
721 cagatgtgat cctccggttg gagaagggag aagagcccgg cggttccggc ggagggtcga
781 tggacaagaa gtacagcatc ggcctggcca tcggcaccaa ctctgtcggc tgggccgtga
841 tcaccgacga gtacaaggtg cccagcaaga aattcaaggt gctgggcaac accgaccggc
901 acagcatcaa gaagaacctg atcggcgccc tgctgttcga cagcggagaa acagccgagg
961 ccacccggct gaagagaacc gccagaagaa gatacaccag acggaagaac cggatctgct
1021 atctgcaaga gatcttcagc aacgagatgg ccaaggtgga cgacagcttc ttccacagac
1081 tggaagagtc cttcctggtg gaagaggata agaagcacga gcggcacccc atcttcggca
1141 acatcgtgga cgaggtggcc taccacgaga agtaccccac catctaccac ctgagaaaga
1201 aactggtgga cagcaccgac aaggccgacc tgcggctgat ctatctggcc ctggcccaca
1261 tgatcaagtt ccggggccac ttcctgatcg agggcgacct gaaccccgac aacagcgacg
1321 tggacaagct gttcatccag ctggtgcaga cctacaacca gctgttcgag gaaaacccca
1381 tcaacgccag cggcgtggac gccaaggcca tcctgtctgc cagactgagc aagagcagac
1441 ggctggaaaa tctgatcgcc cagctgcccg gcgagaagaa gaatggcctg ttcggcaacc
1501 tgattgccct gagcctgggc ctgaccccca acttcaagag caacttcgac ctggccgagg
1561 atgccaaact gcagctgagc aaggacacct acgacgacga cctggacaac ctgctggccc
1621 agatcggcga ccagtacgcc gacctgtttc tggccgccaa gaacctgtcc gacgccatcc
1681 tgctgagcga catcctgaga gtgaacaccg agatcaccaa ggcccccctg agcgcctcta
1741 tgatcaagag atacgacgag caccaccagg acctgaccct gctgaaagct ctcgtgcggc
1801 agcagctgcc tgagaagtac aaagagattt tcttcgacca gagcaagaac ggctacgccg
1861 gctacatcga tggcggagcc agccaggaag agttctacaa gttcatcaag cccatcctgg
1921 aaaagatgga cggcaccgag gaactgctcg tgaagctgaa cagagaggac ctgctgcgga
1981 agcagcggac cttcgacaac ggcagcatcc cccaccagat ccacctggga gagctgcacg
2041 ccattctgcg gcggcaggaa gatttttacc cattcctgaa ggacaaccgg gaaaagatcg
2101 agaagatcct gaccttccgc atcccctact acgtgggccc tctggccagg ggaaacagca
2161 gattcgcctg gatgaccaga aagagcgagg aaaccatcac cccctggaac ttcgaggaag
2221 tggtggacaa gggcgccagc gcccagagct tcatcgagcg gatgaccaac ttcgataaga
2281 acctgcccaa cgagaaggtg ctgcccaagc acagcctgct gtacgagtac ttcaccgtgt
2341 acaacgagct gaccaaagtg aaatacgtga ccgagggaat gagaaagccc gccttcctga
2401 gcggcgagca gaaaaaagcc atcgtggacc tgctgttcaa gaccaaccgg aaagtgaccg
2461 tgaagcagct gaaagaggac tacttcaaga aaatcgagtg cttcgactcc gtggaaatct
2521 ccggcgtgga agatcggttc aacgcctccc tgggcacata ccacgatctg ctgaaaatta
2581 tcaaggacaa ggacttcctg gacaatgagg aaaacgagga cattctggaa gatatcgtgc
2641 tgaccctgac actgtttgag gacagagaga tgatcgagga acggctgaaa acctatgccc
2701 acctgttcga cgacaaagtg atgaagcagc tgaagcggcg gagatacacc ggctggggca
2761 ggctgagccg gaagctgatc aacggcatcc gggacaagca gtccggcaag acaatcctgg
2821 atttcctgaa gtccgacggc ttcgccaaca gaaacttcat gcagctgatc cacgacgaca
2881 gcctgacctt taaagaggac atccagaaag cccaggtgtc cggccagggc gatagcctgc
2941 acgagcacat tgccaatctg gccggcagcc ccgccattaa gaagggcatc ctgcagacag
3001 tgaaggtggt ggacgagctc gtgaaagtga tgggccggca caagcccgag aacatcgtga
3061 tcgaaatggc cagagagaac cagaccaccc agaagggaca gaagaacagc cgcgagagaa
3121 tgaagcggat cgaagagggc atcaaagagc tgggcagcca gatcctgaaa gaacaccccg
3181 tggaaaacac ccagctgcag aacgagaagc tgtacctgta ctacctgcag aatgggcggg
3241 atatgtacgt ggaccaggaa ctggacatca accggctgtc cgactacgat gtggacgcta
3301 tcgtgcctca gagctttctg aaggacgact ccatcgataa caaagtgctg actcggagcg
3361 acaagaaccg gggcaagagc gacaacgtgc cctccgaaga ggtcgtgaag aagatgaaga
3421 actactggcg ccagctgctg aatgccaagc tgattaccca gaggaagttc gacaatctga
3481 ccaaggccga gagaggcggc ctgagcgaac tggataaggc cggcttcatc aagagacagc
3541 tggtggaaac ccggcagatc acaaagcacg tggcacagat cctggactcc cggatgaaca
3601 ctaagtacga cgagaacgac aaactgatcc gggaagtgaa agtgatcacc ctgaagtcca
3661 agctggtgtc cgatttccgg aaggatttcc agttttacaa agtgcgcgag atcaacaact
3721 accaccacgc ccacgacgcc tacctgaacg ccgtcgtggg aaccgccctg atcaaaaagt
3781 accctaagct ggaaagcgag ttcgtgtacg gcgactacaa ggtgtacgac gtgcggaaga
3841 tgatcgccaa gagcgagcag gaaatcggca aggctaccgc caagtacttc ttctacagca
3901 acatcatgaa ctttttcaag accgagatta ccctggccaa cggcgagatc cggaagcggc
3961 ctctgatcga gacaaacggc gaaacaggcg agatcgtgtg ggataagggc cgggactttg
4021 ccaccgtgcg gaaagtgctg tctatgcccc aagtgaatat cgtgaaaaag accgaggtgc
4081 agacaggcgg cttcagcaaa gagtctatcc tgcccaagag gaacagcgac aagctgatcg
4141 ccagaaagaa ggactgggac cctaagaagt acggcggctt cgacagcccc accgtggcct
4201 attctgtgct ggtggtggcc aaagtggaaa agggcaagtc caagaaactg aagagtgtga
4261 aagagctgct ggggatcacc atcatggaaa gaagcagctt cgagaagaat cccatcgact
4321 ttctggaagc caagggctac aaagaagtga aaaaggacct gatcatcaag ctgcctaagt
4381 actccctgtt cgagctggaa aacggccgga agagaatgct ggcctctgcc ggcgaactgc
4441 agaagggaaa cgaactggcc ctgccctcca aatatgtgaa cttcctgtac ctggccagcc
4501 actatgagaa gctgaagggc tcccccgagg ataatgagca gaaacagctg tttgtggaac
4561 agcacaaaca ctacctggac gagatcatcg agcagatcag cgagttctcc aagagagtga
4621 tcctggccga cgctaatctg gacaaggtgc tgagcgccta caacaagcac agagacaagc
4681 ctatcagaga gcaggccgag aatatcatcc acctgtttac cctgaccagt ctgggagccc
4741 ctgccgcctt caagtacttt gacaccacca tcgaccggaa gaggtacacc agcaccaaag
4801 aggtgctgga cgccaccctg atccaccaga gcatcaccgg cctgtacgag acacggatcg
4861 acctgtctca gctgggaggc gacgcctatc cctatgacgt gcccgattat gccagcctgg
4921 gcagcggctc ccccaagaaa aaacgcaagg tggaagatcc taagaaaaag cggaaagtgg
4981 acggcattgg tagtgggagc aacggcagca gcggggatcc ggccgcaaca aacttctctc
5041 tgctgaaaca agccggagat gtcgaagaga atcctggacc gatggccaag cctttgtctc
5101 aagaagaatc caccctcatt gaaagagcaa cggctacaat caacagcatc cccatctctg
5161 aagactacag cgtcgccagc gcagctctct ctagcgacgg ccgcatcttc actggtgtca
5221 atgtatatca ttttactggg ggaccttgtg cagaactcgt ggtgctgggc actgctgctg
5281 ctgcggcagc tggcaacctg acttgtatcg tcgcgatcgg aaatgagaac aggggcatct
5341 tgagcccctg cggacggtgc cgacaggtgc ttctcgatct gcatcctggg atcaaagcca
5401 tagtgaagga cagtgatgga cagccgacgg cagttgggat tcgtgaattg ctgccctctg
5461 gttatgtgtg ggagggctaa gaattcgata tcaagcttat cggtaatcaa cctctggatt
5521 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
5581 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
5641 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
5701 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
5761 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
5821 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
5881 ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt gttgccacct
5941 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
6001 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
6061 cgagtcggat ctccctttgg gccgcctccc cgcgtcgact ttaagaccaa tgacttacaa
6121 ggcagctgta gatcttagcc actttttaaa agaaaagggg ggactggaag ggctaattca
6181 ctcccaacga agacaagatc tgctttttgc ttgtactggg tctctctggt tagaccagat
6241 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
6301 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
6361 cctcagaccc ttttagtcag tgtggaaaat ctctagcagg gcccgtttaa acccgctgat
6421 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
6481 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
6541 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
6601 gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg
6661 aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat
6721 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
6781 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
6841 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
6901 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
6961 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
7021 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
7081 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa
7141 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
7201 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
7261 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
7321 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
7381 ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg
7441 aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt
7501 cggatctgat cagcacgtgt tgacaattaa tcatcggcat agtatatcgg catagtataa
7561 tacgacaagg tgaggaacta aaccatggcc aagttgacca gtgccgttcc ggtgctcacc
7621 gcgcgcgacg tcgccggagc ggtcgagttc tggaccgacc ggctcgggtt ctcccgggac
7681 ttcgtggagg acgacttcgc cggtgtggtc cgggacgacg tgaccctgtt catcagcgcg
7741 gtccaggacc aggtggtgcc ggacaacacc ctggcctggg tgtgggtgcg cggcctggac
7801 gagctgtacg ccgagtggtc ggaggtcgtg tccacgaact tccgggacgc ctccgggccg
7861 gccatgaccg agatcggcga gcagccgtgg gggcgggagt tcgccctgcg cgacccggcc
7921 ggcaactgcg tgcacttcgt ggccgaggag caggactgac acgtgctacg agatttcgat
7981 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
8041 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
8101 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
8161 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
8221 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
8281 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
8341 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
8401 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
8461 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
8521 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
8581 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
8641 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
8701 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
8761 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
8821 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
8881 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
8941 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
9001 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
9061 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
9121 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
9181 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
9241 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
9301 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
9361 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
9421 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
9481 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
9541 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
9601 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
9661 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
9721 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
9781 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
9841 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
9901 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
9961 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
10021 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
10081 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
10141 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
10201 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
10261 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
10321 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
10381 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc
10441 ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc
10501 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa
10561 ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc
10621 ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt
10681 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
10741 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
10801 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
10861 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
10921 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
10981 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
11041 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
11101 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
11161 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
11221 atataagcag cgcgttttgc ctgtactggg tctctctggt tagaccagat ctgagcctgg
11281 gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg
11341 cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc
11401 ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag
11461 ggaaaccaga ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag
11521 gcgaggggcg gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga
11581 gagatgggtg cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa
11641 ttcggttaag gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca
11701 gggagctaga acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac
11761 aaatactggg acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat
11821 ataatacagt agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg
11881 aagctttaga caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg
11941 ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat
12001 aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga
12061 gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga
12121 gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta
12181 ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat
12241 ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa
12301 agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc
12361 accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat
12421 cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc
12481 ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat
12541 aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta
12601 ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata
12661 gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg
12721 aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga
12781 tccattcgat tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac aaatggcagt
12841 attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat
12901 agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat
12961 tcaaaatttt cgggtttatt acagggacag cagagatcca gtttggttaa ttaaggtacc
13021 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
13081 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
13141 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
13201 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
13261 cgaaacaccg gagacggttg taaatgagca cacaaaatac acatgctaaa atattatatt
13321 ctatgacctt tataaaatca accaaaatct tctttttaat aactttagta tcaataatta
13381 gaatttttat gttccttttt gcaaactttt aataaaaatg agcaaaataa aaaaacgcta
13441 gttttagtaa ctcgcgttgt tttcttcacc tttaataata gctactccac cacttgttcc
13501 taagcggtca gctcctgctt caatcatttt ttgagcatct tcaaatgttc taactccacc
13561 agctgcttta actaaagcat tgtctttaac aactgacttc attagtttaa catcttcaaa
13621 tgttgcacct gattttgaaa atcctgttga tgttttaaca aattctaatc cagcttcaac
13681 agctatttca caagctttca tgatttcttc ttttgttaat aaacaatttt ccataataca
13741 tttaacaaca tgtgatccag ctgctttttt tacagctttc atgtcttcta aaactaattc
13801 ataatttttg tcttttaatg caccaatatt taataccata tcaatttctg ttgcaccatc
13861 tttaattgct tcagaaactt cgaatgcttt tgtagctgtt gtgcatgcac ctagaggaaa
13921 acctacaaca tttgttattc ctacatttgt gccttttaat aattctttac aatagcttgt
13981 tcaatatgaa ttaacacaaa ctgttgcaaa atcaaattca attgcttcat cacataattg
14041 tttaatttca gctttcgtag catcttgttt taataatgtg tgatctatat atttgtttag
14101 tttcattttt tctcctatat attcattttt aattttaatt ctttaataat ttcgtctact
14161 ttaactttag cgttttgaac agattcacca acacctataa aataaatttt tagtttaggt
14221 tcagttccac ttgggcgaac agcaaatcat gacttatctt ctaaataaaa ttttagtaag
14281 tcttgtcctg gcatattata cattccatcg atgtagtctt caacattaac aactttaagt
14341 ccagcaattt gagttaaggg tgttgctctc aatgatttca ttaatggttc aatttttaat
14401 ttcttttctt ctggtttaaa attcaagttt aaagtgaaag tgtaatatgc acccatttct
14461 ttaaataaat cttctaaata gtctactaat gttttatttt gttttttata aaatcaagca
14521 gcctctgcta ttaatataga agcttgtatt ccatctttat ctctagctga gtcatcaatt
14581 acatatccat aactttcttc ataagcaaaa acaaaattta atccgttatc ttcttcttta
14641 gcaatttctc tacccattca tttaaatcca gttaaagttt ttacaatatt aactccatat
14701 ttttcatgag cgattctatc acccaaatca cttgttacaa aacttgaata tagagccgga
14761 ttttttggaa tgctatttaa gcgttttaga tttgataatt ttcaatcaat taaaattggt
14821 cctgtttgat ttccatctaa tcttacaaaa tgaccatcat gttttattgc cattccaaat
14881 ctgtcagcat ctgggtcatt cataataata atatctgcat catgtttaat accatattca
14941 agcggtattt ttcatgcagg atcaaattct ggatttggat ttacaacatt tttaaatgtt
15001 tcatcttcaa atgcatgctc ttcaacctca ataacgttat atcctgattc acgtaatatt
15061 tttggggtaa atttagttcc tgttccatta actgcgctaa aaataatttt taaatctttt
15121 ttagcttctt gctctttttt gtacgtctct gttttagagc tagaaatagc aagttaaaat
15181 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgaattcgc
15241 tag
//
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