| 质粒名称: | pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 74009 |
| 存储实验室: | Filippo Del Bene |
|
载体骨架基本信息 |
|
| 载体名称: | pminiTol2 |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 3459 bp |
| 完整质粒大小: | 9668 bp |
| 载体修饰: | -- |
| 载体类型: | CRISPR |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | -- |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | 5x UAS |
| 物种来源: | S. cerevisiae (budding yeast) |
| 插入大小: | 382 bp |
| 基因突变: | -- |
| 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: | pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2 was a gift from Filippo Del Bene (Addgene plasmid # 74009 ; http://n2t.net/addgene:74009 ; RRID:Addgene_74009) |
| 参考文献部分: | 2C-Cas9: a versatile tool for clonal analysis of gene function. Di Donato V, De Santis F, Auer TO, Testa N, Sanchez-Iranzo H, Mercader N, Concordet JP, Del Bene F. Genome Res. 2016 Mar 8. 10.1101/gr.196170.115 PubMed 26957310 |
买家导航
pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2载体质粒基本信息
pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2质粒图谱载体图谱和pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2载体序列质粒序列多克隆位点信息
pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2质粒载体简介
pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2质粒序列载体序列
LOCUS pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2 9671 bp ds-DNA circular SYN 10-APR-2019
DEFINITION Tissue-specific knock-out in zebrafish, Cas9 and GFP driven by a UAS
promoter.
ACCESSION .
VERSION .
KEYWORDS pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9671)
AUTHORS Di Donato V, De Santis F, Auer TO, Testa N, Sanchez-Iranzo H,
Mercader N, Concordet JP, Del Bene F
TITLE 2C-Cas9: a versatile tool for clonal analysis of gene function.
JOURNAL Genome Res. 2016 Mar 8.
PUBMED 26957310
REFERENCE 2 (bases 1 to 9671)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 10, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..9671
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 179..198
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(417..521)
/gene="bla"
/label=AmpR promoter
rep_origin complement(599..1054)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(736..757)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 948..967
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 1180..1202
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 1194..1211
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 1195..1211
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 1218..1237
/label=T7
/note="T7 promoter, forward primer"
promoter 1218..1236
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 1895..1989
/label=5X UAS
/bound_moiety="GAL4"
/note="five tandem copies of the ""ScaI site"" 17-mer
CGGAGTACTGTCCTCCG, an upstream activating sequence (UAS)
that efficiently binds yeast Gal4 (Webster et al., 1988;
Pfeiffer et al., 2010)"
promoter 2008..2246
/label=hsp70 promoter
/note="Drosophila melanogaster hsp70Bb promoter"
primer_bind 2193..2214
/label=pCasper-hs
/note="Drosophila Hsp70 promoter, forward primer"
regulatory 2261..2270
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2279..2299
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 2306..6406
/codon_start=1
/product="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes MGAS15252 Type II CRISPR/Cas system"
/label=Cas9
/note="generates RNA-guided double strand breaks in DNA"
/translation="MDKKYSIGLDIGTNSVGWAVITDDYKVPSKKFKVLGNTDRHSIKK
NLIGALLFGSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLADSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQIYNQLFEENPINASRVDAKAILSARLSKSRRL
ENLIAQLPGEKRNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNSEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGAYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDRGMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGHSLHEQIANLAGSPAIKKGILQTVKIVDELVKVMGH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFIKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
RBS 6319..6327
/label=Shine-Dalgarno sequence
/note="full consensus sequence for ribosome-binding sites
upstream of start codons in E. coli; complementary to a
region in the 3' end of the 16S rRNA (Chen et al., 1994)"
CDS 6422..6442
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 6458..6511
/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 6512..7231
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
primer_bind complement(6557..6578)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(6818..6837)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 7165..7186
/label=EGFP-C
/note="EGFP, forward primer"
promoter complement(7239..7256)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(7285..7304)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 7339..7358
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
promoter complement(8030..8048)
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind complement(8031..8048)
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind complement(8066..8082)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(8066..8082)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(8079..8101)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 8090..8106
/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(8114..8144)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 8159..8180
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(8297..8314)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(8468..9056)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(8548..8567)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(9227..416)
/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"
ORIGIN
1 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
61 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
121 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
181 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
241 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
301 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
361 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
421 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
481 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
541 ccacctgatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggaa
601 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
661 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
721 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
781 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
841 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
901 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
961 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
1021 cccgccgcgc ttaatgcgcc gctacagggc gcgtccattc gccattcagg ctgcgcaact
1081 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
1141 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
1201 cgacggccag tgaattgtaa tacgactcac tatagggcga attgggccca gaggtgtaaa
1261 gtacttgagt aattttactt gattactgta cttaagtatt atttttgggg atttttactt
1321 tacttgagta caattaaaaa tcaatacttt tacttttact taattacatt tttttagaaa
1381 aaaaagtact ttttactcct tacaatttta tttacagtca aaaagtactt attttttgga
1441 gatcacttca ttctattttc ccttgctatt accaaaccaa ttgaattgcg ctgatgccca
1501 gtttaattta aatagatccc atgggtcgac tacgcttaca gctccaaaac tctacggtaa
1561 acctacataa actgctggtt ttcaaatttt aaagaattta agggtttaca ggtttactac
1621 tacacagtga tttactgaca catgtaagtg taaatgagtt gaataagtaa gtaagccata
1681 taccacacat gaaacacata cccagaagtc actggtatat atagccgtcc tccagactcc
1741 cagctcgaga gaccgagaga gggtctcagt ttaagagcta tgctggaaac agcatagcaa
1801 gtttaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt
1861 tggtcgacgg atccaagctt gcatgcctgc aggtcggagt actgtcctcc gagcggagta
1921 ctgtcctccg agcggagtac tgtcctccga gcggagtact gtcctccgag cggagtactg
1981 tcctccgagc ggagactcta gcgagcgccg gagtataaat agaggcgctt cgtctacgga
2041 gcgacaattc aattcaaaca agcaaagtga acacgtcgct aagcgaaagc taagcaaata
2101 aacaagcgca gctgaacaag ctaaacaatc tgcagtaaag tgcaagttaa agtgaatcaa
2161 ttaaaagtaa ccagcaacca agtaaatcaa ctgcaactac tgaaatctgc caagaagtaa
2221 ttattgaata caagaagaga actctgaata gggaattggg gccaccatgg cttctccacc
2281 taagaagaag agaaaggtgg gaagcatgga taagaagtat agcatcggcc tggatattgg
2341 aactaactcc gtgggttggg cagtgattac agacgactac aaggtcccta gcaagaaatt
2401 taaggtgctg ggtaacaccg acaggcacag catcaagaaa aatctgattg gagccctgct
2461 gttcggttct ggagagactg ccgaagcaac acgcctgaaa agaacagcaa gaaggcgcta
2521 taccagaagg aagaatagaa tctgttacct gcaggagatt ttctctaacg aaatggctaa
2581 ggtggacgat tcattctttc ataggctgga ggaaagtttc ctggtcgagg aagataagaa
2641 acacgagcgc catcctatct ttggaaacat tgtggacgag gtcgcctatc acgaaaaata
2701 cccaaccatc tatcatctgc gcaagaaact ggctgactct actgataaag ccgacctgag
2761 actgatctat ctggctctgg cccacatgat taagttcagg ggtcattttc tgatcgaggg
2821 cgatctgaac cccgacaatt ccgatgtgga caagctgttc atccagctgg tccagattta
2881 caatcagctg tttgaggaaa accctattaa tgcttccaga gtggacgcaa aagctatcct
2941 gtcagccagg ctgtccaagt cacgcagact ggagaacctg attgcacagc tgcccggaga
3001 aaagaggaac ggtctgtttg gaaatctgat cgctctgagt ctgggcctga ctcctaactt
3061 caaaagcaat tttgatctgg ctgaggacgc caaactgcag ctgtcaaagg acacatatga
3121 cgatgacctg gataacctgc tggcacagat cggagatcag tacgctgacc tgttcctggc
3181 tgccaaaaat ctgtccgacg caatcctgct gtcagatatt ctgagagtga acagcgagat
3241 tacaaaagct cctctgagtg ccagcatgat caagagatat gacgagcacc atcaggatct
3301 gaccctgctg aaggctctgg tcaggcagca gctgccagag aagtacaagg aaattttctt
3361 tgatcagtcc aagaacggct acgccggtta tatcgacgga ggcgcatcac aggaggaatt
3421 ctacaagttt atcaaaccta ttctggagaa gatggacgga actgaggaac tgctggtgaa
3481 actgaataga gaggacctgc tgaggaagca gcgcacattt gataacggtt ccatcccaca
3541 ccagattcat ctgggagagc tgcacgctat cctgaggcgc caggaagact tctacccctt
3601 tctgaaagat aaccgcgaga agatcgaaaa aattctgacc ttcagaatcc cttactatgt
3661 gggtccactg gctcgcggaa acagcagatt tgcctggatg actcgcaaat ccgaggaaac
3721 cattactcct tggaacttcg aggaagtggt cgataagggc gcctctgcac agtccttcat
3781 cgagagaatg actaattttg acaaaaacct gcccaatgag aaagtgctgc ctaagcactc
3841 cctgctgtac gagtatttca ctgtctataa cgaactgaca aaggtgaaat acgtcaccga
3901 gggcatgaga aagccagcct tcctgtcagg agagcagaag aaagcaatcg tggatctgct
3961 gtttaaaacc aacaggaaag tgactgtcaa gcagctgaag gaggactact tcaagaaaat
4021 tgaatgcttc gattccgtgg agatcagcgg agtcgaagac agatttaacg caagcctggg
4081 cgcttaccac gatctgctga agatcattaa ggataaagac ttcctggaca acgaggaaaa
4141 tgaggatatc ctggaagaca ttgtgctgac actgaccctg tttgaggaca gaggaatgat
4201 cgaggaaaga ctgaaaacct atgctcatct gttcgatgac aaggtgatga aacagctgaa
4261 gagaaggcgc tacactggct ggggtagact gagcaggaag ctgatcaacg gcattaggga
4321 taaacagtca ggaaagacaa tcctggactt tctgaaaagt gatggcttcg ccaaccgcaa
4381 ttttatgcag ctgattcacg atgacagtct gaccttcaaa gaggacatcc agaaggctca
4441 ggtgtctgga cagggccact ccctgcatga gcagattgca aacctggctg gaagcccagc
4501 catcaagaaa ggcattctgc agacagtgaa aatcgtcgat gagctggtga aagtcatggg
4561 ccataagccc gaaaacatcg tgattgagat ggctcgcgaa aatcagacaa cccagaaggg
4621 tcagaagaac agtagagaga ggatgaaaag aatcgaggaa ggcattaagg agctgggtag
4681 ccagatcctg aaagagcacc cagtggaaaa cacacagctg cagaatgaga agctgtatct
4741 gtactatctg cagaatggaa gagatatgta cgtggaccag gagctggata ttaacaggct
4801 gtctgattac gacgtggatc atatcgtccc ccagagtttc atcaaagatg acagcattga
4861 caacaaggtg ctgaccaggt ccgacaaaaa cagaggaaaa tcagataatg tccctagtga
4921 ggaagtggtc aagaaaatga agaactactg gagacagctg ctgaatgcca aactgatcac
4981 tcagaggaag tttgataacc tgacaaaagc agagcgcggt ggactgtcag aactggacaa
5041 agctggattc atcaagaggc agctggtgga aacacgccag atcactaaac acgtcgcaca
5101 gattctggat agtcgcatga acacaaagta cgatgagaat gacaaactga tcagagaagt
5161 gaaggtcatt accctgaaga gtaaactggt cagcgacttt aggaaagatt tccagtttta
5221 taaggtccgc gagattaaca attatcacca tgcccatgac gcatacctga acgccgtggt
5281 cggtaccgca ctgatcaaga aatacccaaa actggagagc gaattcgtgt acggagacta
5341 taaggtgtac gatgtcagaa aaatgatcgc caagtccgag caggaaattg gaaaagctac
5401 tgccaagtat ttcttttact caaacatcat gaatttcttt aagacagaga tcaccctggc
5461 caatggagaa atccgcaaaa ggcccctgat tgagacaaac ggagagacag gcgaaatcgt
5521 gtgggacaaa ggcagagatt ttgcaaccgt gaggaaggtc ctgagcatgc ctcaagtgaa
5581 tatcgtcaag aaaactgagg tgcagacagg cggtttctca aaagaaagta ttctgccaaa
5641 acgcaactct gataagctga tcgctagaaa gaaagactgg gaccctaaga agtatggagg
5701 ctttgactct cccactgtgg catactccgt cctggtggtc gctaaggtgg agaagggcaa
5761 aagcaagaaa ctgaaatctg tcaaggagct gctgggtatc acaattatgg agagaagctc
5821 tttcgagaag aacccaatcg attttctgga ggccaaaggt tataaggaag tgaagaaaga
5881 cctgatcatt aaactgccca agtacagtct gtttgagctg gaaaacggca ggaaacgcat
5941 gctggcaagc gctggagagc tgcagaaagg caatgaactg gccctgcctt ctaagtacgt
6001 gaacttcctg tatctggcaa gccactacga gaagctgaaa ggatctccag aggataacga
6061 acagaaacag ctgtttgtgg agcagcacaa gcattatctg gacgagatca ttgaacagat
6121 tagcgagttc tctaaaagag tgatcctggc cgacgcaaat ctggataagg tcctgtctgc
6181 ttacaacaaa cacagagata agcccatcag ggagcaggcc gaaaatatca ttcatctgtt
6241 cactctgaca aacctgggcg cacctgcagc tttcaagtac ttcgacacta caatcgatag
6301 aaagaggtac acctccacta aggaggtgct ggacgctaca ctgatccatc agagtattac
6361 cggcctgtac gaaacaagga ttgacctgtc tcagctgggt ggcgactcac cggtgagatc
6421 tcctaagaag aagagaaagg tgaggtccgg cggcggagag ggcagaggaa gtcttctaac
6481 atgcggtgac gtggaggaga atcccggccc aatggtgagc aagggcgagg agctgttcac
6541 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
6601 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
6661 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
6721 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
6781 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
6841 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
6901 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
6961 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
7021 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
7081 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa
7141 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
7201 cactctcggc atggacgagc tgtacaagta atctagaact atagtgagtc gtattacgta
7261 gatccagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
7321 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
7381 tgcaataaac cttgtacaaa gtgggggatc ctacgcttac agctccaaaa ctctacggta
7441 aacctacata aactgctggt tttcaaattt taaagaattt aagggtttac aggtttacta
7501 ctacacagtg atttactgac acatgtaagt gtaaatgagt tgaataagta agtaagccat
7561 ataccacaca tgaaacacat acccagaagt cactggtata tatagccgtc ctccagactc
7621 ccagctcgag acgattagat ctaatgcgtc tcgtttaaga gctatgctgg aaacagcata
7681 gcaagtttaa ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt
7741 tttttcaatg atgaattaat tcgagctcca ccgcggaagc ttaaacaaga atctctagtt
7801 ttctttcttg cttttacttt tacttcctta atactcaagt acaattttaa tggagtactt
7861 ttttactttt actcaagtaa gattctagcc agatactttt acttttaatt gagtaaaatt
7921 ttccctaagt acttgtactt tcacttgagt aaaatttttg agtacttttt acacctctgc
7981 tcgaccatat gggagagctc ccaacgcgtt ggatgcatag cttgagtatt ctatagtgtc
8041 acctaaatag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
8101 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
8161 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
8221 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
8281 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
8341 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
8401 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
8461 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
8521 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
8581 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
8641 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
8701 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
8761 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
8821 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
8881 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
8941 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
9001 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
9061 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggca
9121 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
9181 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
9241 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
9301 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
9361 taccgcgaga tccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
9421 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
9481 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
9541 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
9601 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
9661 gtcctccgat c
//
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pUAS:Cas9T2AGFP;U6:sgRNA1;U6:sgRNA2质粒载体应用举例
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