| 质粒名称: | pCI152-pLenti-SiT-Cas12a |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 128405 |
| 存储实验室: | Randall Platt |
|
载体骨架基本信息 |
|
| 载体名称: | pLenti |
| 载体出品公司: | -- |
| 空载体大小: | 4800 bp |
| 完整质粒大小: | 13260 bp |
| 载体修饰: | -- |
| 载体类型: | 哺乳动物表达，慢病毒 |
| 筛选标记: | 嘌呤霉素 |
| 载体抗性: | 氨苄青霉素，100μg/mL |
| 生长温度: | 37 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | -- |
|
插入基因信息 |
|
| 插入基因名称: | AsCas12a-Triplex |
| 物种来源: | -- |
| 插入大小: | 4300 bp |
| 基因突变: | -- |
| 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: | pCI152-pLenti-SiT-Cas12a was a gift from Randall Platt (Addgene plasmid # 128405 ; http://n2t.net/addgene:128405 ; RRID:Addgene_128405) |
| 参考文献部分: | Multiplexed genome engineering by Cas12a and CRISPR arrays encoded on single transcripts. Campa CC, Weisbach NR, Santinha AJ, Incarnato D, Platt RJ. Nat Methods. 2019 Aug 12. pii: 10.1038/s41592-019-0508-6. doi: 10.1038/s41592-019-0508-6. 10.1038/s41592-019-0508-6 PubMed 31406383 |
买家导航
pCI152-pLenti-SiT-Cas12a载体质粒基本信息
pCI152-pLenti-SiT-Cas12a质粒图谱载体图谱和pCI152-pLenti-SiT-Cas12a载体序列质粒序列多克隆位点信息
pCI152-pLenti-SiT-Cas12a质粒载体简介
pCI152-pLenti-SiT-Cas12a质粒序列载体序列
LOCUS pCI152-pLenti-SiT-Cas12a 13286 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Express both AsCas12a and CRISPR arrays on single transcripts.
ACCESSION .
VERSION .
KEYWORDS pCI152-pLenti-SiT-Cas12a
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 13286)
AUTHORS Campa CC, Weisbach NR, Santinha AJ, Incarnato D, Platt RJ
TITLE Multiplexed genome engineering by Cas12a and CRISPR arrays encoded
on single transcripts.
JOURNAL Nat Methods. 2019 Aug 12. pii: 10.1038/s41592-019-0508-6. doi:
10.1038/s41592-019-0508-6.
PUBMED 31406383
REFERENCE 2 (bases 1 to 13286)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..13286
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(53..72)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 244..623
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 625..823
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 841..1021
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 1068..1193
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1686..1919
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 2104..2148
/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 2446..2563
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
polyA_signal 2613..2661
/note="synthetic polyadenylation signal"
misc_feature 2680..3268
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
CDS 2786..2797
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind 3195..3215
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(3567..3647)
/codon_start=1
/product="three tandem HA epitope tags"
/label=3xHA
/translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
CDS complement(3654..4247)
/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 complement(3734..3754)
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
CDS complement(4248..4304)
/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 complement(4320..4367)
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS complement(4368..8288)
/codon_start=1
/product="CRISPR-associated protein from Acidaminococcus
sp. BV3L6 (Zetsche et al., 2015)"
/label=AsCpf1
/translation="MTQFEGFTNLYQVSKTLRFELIPQGKTLKHIQEQGFIEEDKARND
HYKELKPIIDRIYKTYADQCLQLVQLDWENLSAAIDSYRKEKTEETRNALIEEQATYRN
AIHDYFIGRTDNLTDAINKRHAEIYKGLFKAELFNGKVLKQLGTVTTTEHENALLRSFD
KFTTYFSGFYENRKNVFSAEDISTAIPHRIVQDNFPKFKENCHIFTRLITAVPSLREHF
ENVKKAIGIFVSTSIEEVFSFPFYNQLLTQTQIDLYNQLLGGISREAGTEKIKGLNEVL
NLAIQKNDETAHIIASLPHRFIPLFKQILSDRNTLSFILEEFKSDEEVIQSFCKYKTLL
RNENVLETAEALFNELNSIDLTHIFISHKKLETISSALCDHWDTLRNALYERRISELTG
KITKSAKEKVQRSLKHEDINLQEIISAAGKELSEAFKQKTSEILSHAHAALDQPLPTTL
KKQEEKEILKSQLDSLLGLYHLLDWFAVDESNEVDPEFSARLTGIKLEMEPSLSFYNKA
RNYATKKPYSVEKFKLNFQMPTLASGWDVNKEKNNGAILFVKNGLYYLGIMPKQKGRYK
ALSFEPTEKTSEGFDKMYYDYFPDAAKMIPKCSTQLKAVTAHFQTHTTPILLSNNFIEP
LEITKEIYDLNNPEKEPKKFQTAYAKKTGDQKGYREALCKWIDFTRDFLSKYTKTTSID
LSSLRPSSQYKDLGEYYAELNPLLYHISFQRIAEKEIMDAVETGKLYLFQIYNKDFAKG
HHGKPNLHTLYWTGLFSPENLAKTSIKLNGQAELFYRPKSRMKRMAHRLGEKMLNKKLK
DQKTPIPDTLYQELYDYVNHRLSHDLSDEARALLPNVITKEVSHEIIKDRRFTSDKFFF
HVPITLNYQAANSPSKFNQRVNAYLKEHPETPIIGIDRGERNLIYITVIDSTGKILEQR
SLNTIQQFDYQKKLDNREKERVAARQAWSVVGTIKDLKQGYLSQVIHEIVDLMIHYQAV
VVLENLNFGFKSKRTGIAEKAVYQQFEKMLIDKLNCLVLKDYPAEKVGGVLNPYQLTDQ
FTSFAKMGTQSGFLFYVPAPYTSKIDPLTGFVDPFVWKTIKNHESRKHFLEGFDFLHYD
VKTGDFILHFKMNRNLSFQRGLPGFMPAWDIVFEKNETQFDAKGTPFIAGKRIVPVIEN
HRFTGRYRDLYPANELIALLEEKGIVFRDGSNILPKLLENDDSHAIDTMVALIRSVLQM
RNSNAATGEDYINSPVRDLNGVCFDSRFQNPEWPMDADANGAYHIALKGQLLLNHLKES
KDLKLQNGISNQDWLAYIQELRN"
regulatory 4639..4648
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
primer_bind complement(8335..8354)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(8336..8354)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter complement(8399..8602)
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
enhancer 8603..8906
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
LTR 9044..9277
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
primer_bind complement(9303..9320)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 9309..9533
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 9579..10007
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(9666..9685)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 9876..9897
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(10016..10036)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 10021..10350
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 10201..10336
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 10263..10282
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
promoter 10398..10445
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 10464..10838
/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 10968..11089
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(11005..11024)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 11059..11078
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(11138..11154)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(11138..11154)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(11151..11173)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 11162..11178
/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(11186..11216)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 11231..11252
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(11369..11386)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(11540..12128)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(11620..11639)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(12299..13159)
/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 12922..12941
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(13160..13264)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
61 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
121 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
181 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatataccc
241 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
301 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
361 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
421 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
481 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
541 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
601 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
661 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
721 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
781 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagcgcgttt tgcctgtact
841 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
901 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
961 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
1021 agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca
1081 ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc
1141 caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta
1201 agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa
1261 aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc
1321 ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc
1381 ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg
1441 tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc
1501 aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc tggaggagga
1561 gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca
1621 ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg
1681 ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg
1741 tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac
1801 aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc
1861 aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg
1921 gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt
1981 tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga
2041 gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa
2101 gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt
2161 aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta
2221 ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca
2281 ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata
2341 gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa cggatcggca
2401 ctgcgtgcgc caattctgca gacaaatggc agtattcatc cacaatttta aaagaaaagg
2461 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
2521 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
2581 cagcagagat ccagtttggt taattaaggt accacacaaa aaaccaacac acagatctaa
2641 tgaaaataaa gatcttttat tacgcgtcat ggattcgagg cggggaggcg gcccaaaggg
2701 agatccgact cgtctgaggg cgaaggcgaa gacgcggaag aggccgcaga gccggcagca
2761 ggccgcggga aggaaggtcc gctggattga gggccgaagg gacgtagcag aaggacgtcc
2821 cgcgcagaat ccaggtggca acacaggcga gcagccaagg aaaggacgat gatttccccg
2881 acaacaccac ggaattgtca gtgcccaaca gccgagcccc tgtccagcag cgggcaaggc
2941 aggcggcgat gagttccgcc gtggcaatag ggagggggaa agcgaaagtc ccggaaagga
3001 gctgacaggt ggtggcaatg ccccaaccag tgggggttgc gtcagcaaac acagtgcaca
3061 ccacgccacg ttgcctgaca acgggccaca actcctcata aagagacagc aaccaggatt
3121 tatacaagga ggagaaaatg aaagccatac gggaagcaat agcatgatac aaaggcatta
3181 aagcagcgta tccacatagc gtaaaaggag caacatagtt aagaatacca gtcaatcttt
3241 cacaaatttt gtaatccaga ggttgattgt cgacttaacg cgatctatgt ctagaatcta
3301 caagagtaga aattagagac gagaaatcat gaaagcttgt gaaatagctg ttgaagctgg
3361 attagaattt gcgtctccat ctacaagagt agaaatttcg cagaccaata ccaggatctt
3421 gccatcctat ggaactgcct cggtgagttt tgcttttttt tttttaaagc tagggaaagg
3481 ccaaaaagca aaacctgaga aaacaaaagg ttgttttctc aggaaaagaa aaacctttac
3541 aaccctactg acgaatcgaa ttcttaggca tagtcgggga catcatatgg gtatgcataa
3601 tcaggcacgt cgtagggata agcgtaatct ggaacatcgt atgggtagga tccggcaccg
3661 ggcttgcggg tcatgcacca ggtgcgcggt ccttcgggca cctcgacgtc ggcggtgacg
3721 gtgaagccga gccgctcgta gaaggggagg ttgcggggcg cggaggtctc caggaaggcg
3781 ggcaccccgg cgcgctcggc cgcctccact ccggggagca cgacggcgct gcccagaccc
3841 ttgccctggt ggtcgggcga gactccgacg gtggccagga accacgcggg ctccttgggc
3901 cggtgcggcg ccaggaggcc ttccatctgt tgctgcgcgg ccagccggga accgctcaac
3961 tcggccatgc gcgggccgat ctcggcgaac accgcccccg cttcgacgct ctccggcgtg
4021 gtccagaccg ccacggcggc gccgtcgtcc gcgacccaca ccttgccgat gtcgagcccg
4081 acgcgcgtga ggaagagttc ttgcagctcg gtgacccgct cgatgtggcg gtccggatcg
4141 acggtgtggc gcgtggcggg gtagtcggcg aacgcggcgg cgagggtgcg tacggccctg
4201 gggacgtcgt cgcgggtggc gaggcgcacc gtgggcttgt actcggttgg cccgggattc
4261 tcttcgacat cccctgcttg tttcaacagg gagaagttag tggctccgct tccggatccc
4321 tttttctttt ttgcctggcc ggcctttttc gtggccgccg gccttttgtt gcgcagctcc
4381 tggatgtagg ccagccagtc ctgattggag atgccgttct gcagcttcag atccttgctc
4441 tccttcaggt gattcagcag cagctggccc ttcagggcga tgtggtaggc gccattggca
4501 tcggcgtcca tgggccactc tgggttctga aaccgggagt cgaagcacac gccattcaga
4561 tcgcgcacgg ggctgttgat atagtcctcg cctgtggcgg cattggagtt ccgcatctgc
4621 agcacgctgc ggatcagggc caccatggtg tcgatggcgt gagaatcgtc attctccagc
4681 agctttggca ggatgttgga gccatccctg aacacgatgc ccttctcctc cagcagggcg
4741 atcagctcgt tggcaggata caggtcccgg tatctgccgg tgaatctgtg attctcgatc
4801 actggcacga ttctcttgcc ggcgatgaaa ggggtgccct tggcgtcaaa ctgtgtctcg
4861 ttcttctcga acacgatatc ccatgcaggc ataaagccgg gcaggcccct ctggaaggac
4921 agatttctgt tcatcttaaa gtgcaggatg aagtcgccgg ttttcacgtc gtagtgcaga
4981 aagtcgaagc cctccaggaa gtgcttgcgg ctctcgtgat tcttgatggt tttccacacg
5041 aaggggtcca cgaagccggt caggggatcg atcttagatg tatatggggc aggcacgtaa
5101 aacaggaagc cagactgggt gcccatcttg gcaaaggagg tgaactggtc tgtcagctgg
5161 tatgggttca gcacgcctcc cactttctct gctggatagt ccttcagcac caggcaattc
5221 agcttatcga tcagcatctt ctcgaactgc tggtacacgg ccttctcggc gatgccggtc
5281 ctcttgctct taaagccgaa attcaggttc tccagcacca ccacggcctg gtagtggatc
5341 atcaggtcca cgatctcgtg gatgacctgg ctcagatagc cctgcttcag atccttgatt
5401 gtgcccacca cagaccaggc ctgccttgct gccaccctct ccttctccct gttgtccagc
5461 ttcttctggt aatcaaactg ctggatggtg ttcaggctcc gctgctccag gatcttgccg
5521 gtggagtcga tcactgtgat atagatcagg tttctctcgc cccgatcgat gccgatgata
5581 ggtgtctcgg ggtgctcctt caggtaggca ttcaccctct ggttgaactt agatggggaa
5641 ttggcggcct gatagttcag tgtgataggc acgtggaaaa agaacttgtc gctggtaaag
5701 cgcctatcct tgatgatctc gtgagacacc tccttggtga tcacgttggg cagcagggcc
5761 ctggcctcat cagacaggtc gtgggacagt ctgtgattca catagtcgta cagctcctgg
5821 tacagggtgt cggggattgg ggttttctga tccttcagct tcttgttcag catcttctct
5881 cccagccggt gtgccatcct cttcatcctg gacttagggc ggtagaacag ctcggcctgg
5941 ccattcagct tgatgcttgt cttggccagg ttctctggag aaaacaggcc ggtccaatac
6001 agtgtgtgca gattaggctt gccgtggtgg cccttggcaa agtccttgtt atagatctgg
6061 aacaggtaca gcttgcctgt ctccacggca tccatgatct ccttctcggc gattctctgg
6121 aagctgatgt ggtacagcag gggattcagc tcggcatagt actcgcccag gtccttatac
6181 tgagaggatg gccgcaggct agacagatcg atagaggttg tcttggtata cttggacaga
6241 aaatcccttg tgaagtcgat ccacttgcac agggcctctc tgtagccctt ctggtcgccg
6301 gttttcttgg cgtaggctgt ctgaaacttc tttggctcct tctcaggatt gttcaggtcg
6361 tagatctcct ttgtgatctc cagaggctcg atgaaattgt tggacagcag gatgggggtt
6421 gtgtgggtct gaaagtgggc tgtcacggcc ttcagctggg tgctgcactt tgggatcatc
6481 ttggcggcat cagggaagta gtcatagtac atcttatcaa agccctcgct ggttttctct
6541 gtgggctcga agctcagggc cttatacctg cccttctgct ttggcatgat gcccagatag
6601 tacaggccgt tcttcacaaa caggatggcg ccattgttct tctccttatt cacgtcccag
6661 ccagaggcca gtgtaggcat ctgaaagttc agcttgaact tctccacgga gtagggcttc
6721 ttggtggcat aatttctggc cttgttgtag aagctcagag aaggctccat ctccagcttg
6781 atgccggtca gccgggcaga gaactcgggg tccacctcgt tggactcatc cacggcaaac
6841 cagtccagca ggtggtacag gcccagcagg ctgtccagct gagacttcag gatctccttc
6901 tcctcctgct tcttcagggt tgtaggcagt ggctgatcca gggcggcgtg tgcgtgggac
6961 aggatctcgc tggttttctg cttgaaggcc tcgctcagct ccttgcctgc ggcagagatg
7021 atctcctgca ggttgatatc ctcgtgcttc aggctgcgct gcaccttctc cttggcagac
7081 ttggtgatct tgcctgtcag ctcggagatt ctccgctcat acagggcatt cctcagtgta
7141 tcccagtggt cgcacagggc gctgctgatt gtctccagct tcttgtggct gatgaagatg
7201 tgtgtcaggt cgatgctgtt cagctcgtta aacagggcct cggctgtctc cagcacgttc
7261 tcgtttctca gcagtgtctt gtacttgcag aaggactgga tcacttcctc gtcgctctta
7321 aactcctcca ggatgaaaga cagggtgttc ctatcggaca ggatctgctt aaacaggggg
7381 atgaatctgt gtggcaggga ggcgatgatg tgggctgtct catcattctt ctggatggcc
7441 agattcagca cctcgttcag gcccttgatc ttctcggtgc ctgcctcccg agagattcct
7501 cccagcagct ggttatacag gtcgatctgg gtctgtgtca gcagctggtt ataaaaaggg
7561 aaggaaaaca cctcctcgat ggaggtgctc acgaagatgc cgatggcctt cttcacgttc
7621 tcaaagtgct cccgcaggct gggcacggcg gtgatcaggc gtgtgaagat gtgacaattc
7681 tccttaaact tggggaagtt gtcctgcacg atgcggtgtg ggatggctgt gctgatatcc
7741 tcggcgctga acacgttctt cctgttctca taaaagccgg agaagtaggt tgtaaacttg
7801 tcgaagctcc gcagcagggc gttctcgtgc tcggttgtgg tcacggtgcc cagctgcttc
7861 agcaccttgc cattaaacag ctcggccttg aacaggccct tgtagatctc ggcgtgtctc
7921 ttattgatgg catcggtcag gttgtctgtc cggccgatga agtagtcgtg gatggcattg
7981 cgatatgtgg cctgctcctc gatcagggcg ttccttgtct cctcggtttt ctcctttcta
8041 taggagtcga tggcggcgct caggttctcc caatccagct gcaccagctg caggcactgg
8101 tcggcatagg tcttgtagat ccgatcgatg atgggcttca gctccttgta gtgatcattg
8161 cgggccttgt cctcctcgat gaagccctgc tcctggatgt gcttcagggt cttgccctgt
8221 gggatcagct caaaccgcag tgtcttgctc acctgataca ggttggtaaa gccctcgaac
8281 tgtgtcatgg tggcggtacc aagcttaagt ttaaacgcta gccagcttgg gtctccctat
8341 agtgagtcgt attaatttcg ataagccagt aagcagtggg ttctctagtt agccagagag
8401 ctctgcttat atagacctcc caccgtacac gcctaccgcc catttgcgtc aatggggcgg
8461 agttgttacg acattttgga aagtcccgtt gattttggtg ccaaaacaaa ctcccattga
8521 cgtcaatggg gtggagactt ggaaatcccc gtgagtcaaa ccgctatcca cgcccattga
8581 tgtactgcca aaaccgcatc accatggtaa tagcgatgac taatacgtag atgtactgcc
8641 aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc catttaccgt
8701 cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact gccaagtggg
8761 cagtttaccg taaatactcc acccattgac gtcaatggaa agtccctatt ggcgttacta
8821 tgggaacata cgtcattatt gacgtcaatg ggcgggggtc gttgggcggt cagccaggcg
8881 ggccatttac cgtaagttat gtaacgcgga actccatata tgggctatga actaatgacc
8941 ccgtaattga ttactattaa taactagtca cgctcgagtt taagaccaat gacttacaag
9001 gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac
9061 tcccaacgaa gacaagatct gctttttgct tgtactgggt ctctctggtt agaccagatc
9121 tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg
9181 ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc
9241 ctcagaccct tttagtcagt gtggaaaatc tctagcaggg cccgtttaaa cccgctgatc
9301 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
9361 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
9421 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
9481 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
9541 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
9601 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
9661 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
9721 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
9781 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
9841 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
9901 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
9961 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
10021 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
10081 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
10141 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
10201 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
10261 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
10321 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
10381 ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc atagtataat
10441 acgacaaggt gaggaactaa accatggcca agttgaccag tgccgttccg gtgctcaccg
10501 cgcgcgacgt cgccggagcg gtcgagttct ggaccgaccg gctcgggttc tcccgggact
10561 tcgtggagga cgacttcgcc ggtgtggtcc gggacgacgt gaccctgttc atcagcgcgg
10621 tccaggacca ggtggtgccg gacaacaccc tggcctgggt gtgggtgcgc ggcctggacg
10681 agctgtacgc cgagtggtcg gaggtcgtgt ccacgaactt ccgggacgcc tccgggccgg
10741 ccatgaccga gatcggcgag cagccgtggg ggcgggagtt cgccctgcgc gacccggccg
10801 gcaactgcgt gcacttcgtg gccgaggagc aggactgaca cgtgctacga gatttcgatt
10861 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
10921 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
10981 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
11041 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
11101 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
11161 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
11221 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
11281 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
11341 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
11401 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
11461 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
11521 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
11581 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
11641 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
11701 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
11761 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
11821 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
11881 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
11941 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
12001 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
12061 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
12121 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
12181 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
12241 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
12301 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
12361 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
12421 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
12481 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
12541 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
12601 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
12661 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
12721 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
12781 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
12841 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
12901 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
12961 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
13021 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
13081 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
13141 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
13201 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
13261 cgcgcacatt tccccgaaaa gtgcca
//
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