| 质粒名称: | pICH86966::AtU6p::sgRNA_PDS |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 46966 |
| 存储实验室: | Sophien Kamoun |
|
载体骨架基本信息 |
|
| 载体名称: | pICH86966 |
| 载体出品公司: | -- |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | 6340 bp |
| 完整质粒大小: | 6552 bp |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 低拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | AtU6p::sgRNA_PDS |
| 物种来源: | -- |
| 插入大小: | 212 bp |
| 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: | AgeI(未破坏) |
| 3' 克隆位点: | NotI (未破坏) |
| 5' 测序引物 | -- |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pICH86966::AtU6p::sgRNA_PDS was a gift from Sophien Kamoun (Addgene plasmid # 46966 ; http://n2t.net/addgene:46966 ; RRID:Addgene_46966) |
| 参考文献部分: | Targeted mutagenesis in the model plant Nicotiana benthamiana using Cas9 RNA-guided endonuclease. Nekrasov V, Staskawicz B, Weigel D, Jones JD, Kamoun S. Nat Biotechnol. 2013 Aug 8;31(8):691-3. doi: 10.1038/nbt.2655. 10.1038/nbt.2655 PubMed 23929340 |
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
载体简介
载体序列
LOCUS pICH86966::AtU6p::sgRNA_PDS 6551 bp ds-DNA circular SYN 19-APR-2019
DEFINITION Expresses an sgRNA targeting the PDS gene in Nicotiana benthamiana
from the Arabidopsis U6 promoter.
ACCESSION .
VERSION .
KEYWORDS pICH86966::AtU6p::sgRNA_PDS
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6551)
AUTHORS Nekrasov V, Staskawicz B, Weigel D, Jones JD, Kamoun S
TITLE Targeted mutagenesis in the model plant Nicotiana benthamiana using
Cas9 RNA-guided endonuclease.
JOURNAL Nat Biotechnol. 2013 Aug 8;31(8):691-3. doi: 10.1038/nbt.2655.
PUBMED 23929340
REFERENCE 2 (bases 1 to 6551)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 19, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6551
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_feature 32..56
/label=RB T-DNA repeat
/note="right border repeat from nopaline C58 T-DNA"
rep_origin complement(179..767)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(259..278)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(938..1732)
/codon_start=1
/gene="aphA-3"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin"
/translation="MAKMRISPELKKLIEKYRCAKDTEGMSPAKVYKLVGENENLYLKM
TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
promoter complement(1733..1837)
/gene="bla"
/label=AmpR promoter
rep_origin 2120..2314
/label=pVS1 oriV
/note="origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
CDS complement(2380..3453)
/codon_start=1
/product="replication protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 RepA
/translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
QAAADRIRKESRQPPAAGASSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
LVMRYRNLIEGEASAGS"
CDS complement(3882..4511)
/codon_start=1
/product="stability protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 StaA
/translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
misc_feature 4775..4799
/label=LB T-DNA repeat
/note="left border repeat from nopaline C58 T-DNA"
promoter 4977..5156
/label=NOS promoter
/note="nopaline synthase promoter"
promoter 5009..5192
/function="NOS promoter"
/label=NOS promoter
/note="nopaline synthase promoter"
CDS 5213..6007
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(5267..5286)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 5877..5896
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
terminator 6033..6285
/label=NOS terminator
/note="nopaline synthase terminator and poly(A) signal"
promoter 6312..6386
/label=AtU6 promoter
/note="consensus promoter from the three Arabidopsis U6
snRNA genes (Waibel and Filipowicz, 1990; Nekrasov et al.,
2013)"
misc_RNA 6407..6482
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
protein_bind complement(6494..6518)
/gene="mutant version of attB"
/label=attB2
/bound_moiety="BP Clonase(TM)"
/note="recombination site for the Gateway(R) BP reaction"
ORIGIN
1 gggacacgaa gtggtttaaa ctatcagtgt ttgacaggat atattggcgg gtaaacctaa
61 gagaaaagag cgtttattag aataatcgga tatttaaaag ggcgtgaaaa ggtttatccg
121 ttcgtccatt tgtatgtgcc aaccacagga accgtaaaaa ggccgcgttg ctggcgtttt
181 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
241 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
301 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
361 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
421 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
481 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
541 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
601 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
661 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
721 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
781 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
841 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
901 caatctaaag tatatatgag taaacttggt ctgacagcta aaacaattca tccagtaaaa
961 tataatattt tattttctcc caatcaggct tgatccccag taagtcaaaa aatagctcga
1021 catactgttc ttccccgata tcctccctga tcgaccggac gcagaaggca atgtcatacc
1081 acttgtccgc cctgccgctt ctcccaagat caataaagcc acttactttg ccatctttca
1141 caaagatgtt gctgtctccc aggtcgccgt gggaaaagac aagttcctct tcgggctttt
1201 ccgtctttaa aaaatcatac agctcgcgcg gatctttaaa tggagtgtcc tcttcccagt
1261 tttcgcaatc cacatcggcc agatcgttat tcagtaagta atccaattcg gctaagcggc
1321 tgtctaagct attcgtatag ggacaatccg atatgtcgat ggagtgaaag agcctgatgc
1381 actccgcata cagctcgata atcttttcag ggctttgttc atcttcatac tcttccgagc
1441 aaaggacgcc atcggcctca ctcatgagca gattgctcca gccatcatgc cgttcaaagt
1501 gcaggacctt tggaacaggc agctttcctt ccagccatag catcatgtcc ttttcccgtt
1561 ccacatcata ggtggtccct ttataccggc tgtccgtcat ttttaaatat aggttttcat
1621 tttctcccac cagcttatat accttagcag gagacattcc ttccgtatct tttgcgcagc
1681 ggtatttttc gatcagtttt ttcaattccg gtgatattct cattttagcc atactcttcc
1741 tttttcaata ttattgaagc atttatcaag gttattgtct catgagcgga tacatatttg
1801 aatgtattta gaaaaataaa caaatagggg ttccgcgcac gaattggcca gcgctgccat
1861 ttttggggtg aggccgttcg cggccgaggg gcgcagcccc tggggggatg ggagctcggt
1921 gccttgatgt gggcgccggc ggtcgagtgg cgacggcgcg gcttgtccgc gccctggtag
1981 attgcctggc cgtaggccag ccatttttga gcggccagcg gccgcgatag gccgacgcga
2041 agcggcgggg cgtagggagc gcagcgaccg aagggtaggc gctttttgca gctcttcggc
2101 tgtgcgctgg ccagacagtt atgcacaggc caggcgggtt ttaagagttt taataagttt
2161 taaagagttt taggcggaaa aatcgccttt tttctctttt atatcagtca cttacatgtg
2221 tgaccggttc ccaatgtacg gctttgggtt cccaatgtac gggttccggt tcccaatgta
2281 cggctttggg ttcccaatgt acgtgctatc cacaggaaag aagctttttc gacctttttc
2341 ccctgctagg gcaatttgcc ctagcatctg ctccgtacat taggaaccgg cggatgcttc
2401 gccctcgatc aggttgcggt agcgcatgac taggatcggg ccagcctgcc ccgcctcctc
2461 cttcaaatcg tactccggca ggtcatttga cccgatcagc ttgcgcacgg tgaaacagaa
2521 cttcttgaac tctccggcgc tgccactgcg ttcgtagatc gtcttgaaca accatctggc
2581 ttctgccttg cctgcggcgc ggcgtgccag gcggtagaga aaacggccga tgccgggatc
2641 gatcaaaaag taatcggggt gaaccgtcag cacgtccggg ttcttgcctt ctgtgatctc
2701 gcggtacatc caatcagcta gctcgatctc gatgtactcc ggccgcccgg tttcgctctt
2761 tacgatcttg tagcggctaa tcaaggcttc accctcggat accgtcacca ggcggccgtt
2821 cttggccttc ttcgtacgct gcatggcaac gtgcgtggtg tttaaccgaa tgcaggtttc
2881 taccaggtcg tctttctgct ttccgccatc ggctcgccgg cagaacttga gtacgtccgc
2941 aacgtgtgga cggaacacgc ggccgggctt gtctcccttc ccttcccggt atcggttcat
3001 ggattcggtt agatgggaaa ccgccatcag taccaggtcg taatcccaca cactggccat
3061 gccggccggc cctgcggaaa cctctacgtg cccgtctgga agctcgtagc ggatcacctc
3121 gccagctcgt cggtcacgct tcgacagacg gaaaacggcc acgtccatga tgctgcgact
3181 atcgcgggtg cccacgtcat agagcatcgg aacgaaaaaa tctggttgct cgtcgccctt
3241 gggcggcttc ctaatcgacg acgcaccggc tgccggcggt tgccgggatt ctttgcggat
3301 tcgatcagcg gccgcttgcc acgattcacc ggggcgtgct tctgcctcga tgcgttgccg
3361 ctgggcggcc tgcgcggcct tcaacttctc caccaggtca tcacccagcg ccgcgccgat
3421 ttgtaccggg ccggatggtt tgcgaccgct cacgccgatt cctcgggctt gggggttcca
3481 gtgccattgc agggccggca gacaacccag ccgcttacgc ctggccaacc gcccgttcct
3541 ccacacatgg ggcattccac ggcgtcggtg cctggttgtt cttgattttc catgccgcct
3601 cctttagccg ctaaaattca tctactcatt tattcatttg ctcatttact ctggtagctg
3661 cgcgatgtat tcagatagca gctcggtaat ggtcttgcct tggcgtaccg cgtacatctt
3721 cagcttggtg tgatcctccg ccggcaactg aaagttgacc cgcttcatgg ctggcgtgtc
3781 tgccaggctg gccaacgttg cagccttgct gctgcgtgcg ctcggacggc cggcacttag
3841 cgtgtttgtg cttttgctca ttttctcttt acctcattaa ctcaaatgag ttttgattta
3901 atttcagcgg ccagcgcctg gacctcgcgg gcagcgtcgc cctcgggttc tgattcaaga
3961 acggttgtgc cggcggcggc agtgcctggg tagctcacgc gctgcgtgat acgggactca
4021 agaatgggca gctcgtaccc ggccagcgcc tcggcaacct caccgccgat gcgcgtgcct
4081 ttgatcgccc gcgacacgac aaaggccgct tgtagccttc catccgtgac ctcaatgcgc
4141 tgcttaacca gctccaccag gtcggcggtg gcccatatgt cgtaagggct tggctgcacc
4201 ggaatcagca cgaagtcggc tgccttgatc gcggacacag ccaagtccgc cgcctggggc
4261 gctccgtcga tcactacgaa gtcgcgccgg ccgatggcct tcacgtcgcg gtcaatcgtc
4321 gggcggtcga tgccgacaac ggttagcggt tgatcttccc gcacggccgc ccaatcgcgg
4381 gcactgccct ggggatcgga atcgactaac agaacatcgg ccccggcgag ttgcagggcg
4441 cgggctagat gggttgcgat ggtcgtcttg cctgacccgc ctttctggtt aagtacagcg
4501 ataaccttca tgcgttcccc ttgcgtattt gtttatttac tcatcgcatc atatacgcag
4561 cgaccgcatg acgcaagctg ttttactcaa atacacatca cctttttaga cggcggcgct
4621 cggtttcttc agcggccaag ctggccggcc aggccgccag cttggcatca gacaaaccgg
4681 ccaggatttc atgcagccgc acggctgatg ggctgcctgt atcgagtggt gattttgtgc
4741 cgagctgccg gtcggggagc tgttggctgg ctggtggcag gatatattgt ggtgtaaaca
4801 aattgacgct tagacaactt aataacacat tgcggacgtt tttaatgtac tggggtggat
4861 gcagtgggcc ccactctgtg tgccgaattc ggatccggag agcggagaat taagggagtc
4921 acgttatgac ccccgccgat gacgcgggac aagccgtttt acgtttggaa ctgacagaac
4981 cgcaacgttg aaggagccac tcagccgcgg gtttctggag tttaatgagc taagcacata
5041 cgtcagaaac cattattgcg cgttcaaaag tcgcctaagg tcactatcag ctagcaaata
5101 tttcttgtca aaaatgctcc actgacgttc cataaattcc cctcggtatc caattagagt
5161 ctcatattca ctctcaatcc aaataatctg caccggatct ggatcgtttc gcatgattga
5221 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
5281 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
5341 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tccaggacga
5401 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
5461 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
5521 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
5581 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
5641 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
5701 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgtatgcccg acggcgagga
5761 tctcgtcgtg actcatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
5821 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
5881 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
5941 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
6001 cttctgagcg ggtctggatc tagagtcaag cagatcgttc aaacatttgg caataaagtt
6061 tcttaagatt gaatcctgtt gccggtcttg cgatgattat catataattt ctgttgaatt
6121 acgttaagca tgtaataatt aacatgtaat gcatgacgtt atttatgaga tgggttttta
6181 tgattagagt cccgcaatta tacatttaat acgcgataga aaacaaaata tagcgcgcaa
6241 actaggataa attatcgcgc gcggtgtcat ctatgttact agatcgacgc tgcaagaatt
6301 caagcttgga gtgatcaaaa gtcccacatc gatcaggtga tatatagcag cttagtttat
6361 ataatgatag agtcgacata gcgattgccg ttaatttgag agtccagttt tagagctaga
6421 aatagcaagt taaaataagg ctagtccgtt atcaacttga aaaagtggca ccgagtcggt
6481 gctttttttc tagacccagc tttcttgtac aaagttggca ttacgctact agaggatgca
6541 catgtgaccg a
//
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