| 质粒名称: | pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 61594 |
| 存储实验室: | Feng Zhang |
|
载体骨架基本信息 |
|
| 载体名称: | pAAV |
| 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，AAV，CRISPR |
| 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: | hSaCas9 |
| 物种来源: | -- |
| 插入大小: | 3345 bp |
| 基因突变: | D10A, N580A |
| GenBank编号 | CCK74173.1 |
| 启动子 | -- |
| 标签/融合蛋白: |
NLS（插入件上的N端子）
NLS（插接件上的C端子）
3xHA（插入件上的C端子）
|
| 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: | pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA was a gift from Feng Zhang (Addgene plasmid # 61594 ; http://n2t.net/addgene:61594 ; RRID:Addgene_61594) |
| 参考文献部分: | In vivo genome editing using Staphylococcus aureus Cas9. Ran FA, Cong L, Yan WX, Scott DA, Gootenberg JS, Kriz AJ, Zetsche B, Shalem O, Wu X, Makarova KS, Koonin EV, Sharp PA, Zhang F. Nature. 2015 Apr 1. doi: 10.1038/nature14299. 10.1038/nature14299 PubMed 25830891 |
pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA
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pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA载体质粒基本信息
pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA质粒图谱载体图谱和pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA载体序列质粒序列多克隆位点信息
-NLS-3xHA-bGHpA质粒图谱.png)
pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA质粒载体简介
pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA质粒序列载体序列
LOCUS pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA 7090 bp ds-DNA circular SYN 23-AUG-2018
DEFINITION A catalytically inactive SaCas9 (dSaCas9) with the D10A and N580A
mutations. .
ACCESSION .
VERSION .
KEYWORDS pX603-AAV-CMV::NLS-dSaCas9(D10A,N580A)-NLS-3xHA-bGHpA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7090)
AUTHORS Ran FA, Cong L, Yan WX, Scott DA, Gootenberg JS, Kriz AJ, Zetsche B,
Shalem O, Wu X, Makarova KS, Koonin EV, Sharp PA, Zhang F
TITLE In vivo genome editing using Staphylococcus aureus Cas9.
JOURNAL Nature. 2015 Apr 1. doi: 10.1038/nature14299.
PUBMED 25830891
REFERENCE 2 (bases 1 to 7090)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 23, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7090
/organism="synthetic DNA construct"
/mol_type="other DNA"
repeat_region 42..182
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 53..182
/label=AAV2 ITR
enhancer 202..581
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 582..785
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 736..756
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
regulatory 804..813
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 816..836
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 4017..4064
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS 4071..4151
/codon_start=1
/product="three tandem HA epitope tags"
/label=3xHA
/translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
primer_bind complement(4179..4196)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 4185..4392
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
repeat_region 4405..4534
/label=AAV2 ITR (alternate)
/note="Functional equivalent of wild-type AAV2 ITR"
rep_origin complement(4596..5184)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(4676..4695)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5355..6215)
/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 5978..5997
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(6216..6320)
/gene="bla"
/label=AmpR promoter
primer_bind 6388..6406
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind complement(6444..6466)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 6566..6585
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
rep_origin complement(6602..7057)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(6739..6760)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 6951..6970
/label=F1ori-R
/note="F1 origin, reverse primer"
ORIGIN
1 accgcacaga tgcgtaagga gaaaataccg catcaggcgc ccctgcaggc agctgcgcgc
61 tcgctcgctc actgaggccg cccgggcaaa gcccgggcgt cgggcgacct ttggtcgccc
121 ggcctcagtg agcgagcgag cgcgcagaga gggagtggcc aactccatca ctaggggttc
181 ctgcggccgc ctcgaggcgt tgacattgat tattgactag ttattaatag taatcaatta
241 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
301 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
361 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
421 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
481 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
541 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
601 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
661 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
721 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
781 gagctctctg gctaactacc ggtgccacca tggccccaaa gaagaagcgg aaggtcggta
841 tccacggagt cccagcagcc aagcggaact acatcctggg cctggccatc ggcatcacca
901 gcgtgggcta cggcatcatc gactacgaga cacgggacgt gatcgatgcc ggcgtgcggc
961 tgttcaaaga ggccaacgtg gaaaacaacg agggcaggcg gagcaagaga ggcgccagaa
1021 ggctgaagcg gcggaggcgg catagaatcc agagagtgaa gaagctgctg ttcgactaca
1081 acctgctgac cgaccacagc gagctgagcg gcatcaaccc ctacgaggcc agagtgaagg
1141 gcctgagcca gaagctgagc gaggaagagt tctctgccgc cctgctgcac ctggccaaga
1201 gaagaggcgt gcacaacgtg aacgaggtgg aagaggacac cggcaacgag ctgtccacca
1261 aagagcagat cagccggaac agcaaggccc tggaagagaa atacgtggcc gaactgcagc
1321 tggaacggct gaagaaagac ggcgaagtgc ggggcagcat caacagattc aagaccagcg
1381 actacgtgaa agaagccaaa cagctgctga aggtgcagaa ggcctaccac cagctggacc
1441 agagcttcat cgacacctac atcgacctgc tggaaacccg gcggacctac tatgagggac
1501 ctggcgaggg cagccccttc ggctggaagg acatcaaaga atggtacgag atgctgatgg
1561 gccactgcac ctacttcccc gaggaactgc ggagcgtgaa gtacgcctac aacgccgacc
1621 tgtacaacgc cctgaacgac ctgaacaatc tcgtgatcac cagggacgag aacgagaagc
1681 tggaatatta cgagaagttc cagatcatcg agaacgtgtt caagcagaag aagaagccca
1741 ccctgaagca gatcgccaaa gaaatcctcg tgaacgaaga ggatattaag ggctacagag
1801 tgaccagcac cggcaagccc gagttcacca acctgaaggt gtaccacgac atcaaggaca
1861 ttaccgcccg gaaagagatt attgagaacg ccgagctgct ggatcagatt gccaagatcc
1921 tgaccatcta ccagagcagc gaggacatcc aggaagaact gaccaatctg aactccgagc
1981 tgacccagga agagatcgag cagatctcta atctgaaggg ctataccggc acccacaacc
2041 tgagcctgaa ggccatcaac ctgatcctgg acgagctgtg gcacaccaac gacaaccaga
2101 tcgctatctt caaccggctg aagctggtgc ccaagaaggt ggacctgtcc cagcagaaag
2161 agatccccac caccctggtg gacgacttca tcctgagccc cgtcgtgaag agaagcttca
2221 tccagagcat caaagtgatc aacgccatca tcaagaagta cggcctgccc aacgacatca
2281 ttatcgagct ggcccgcgag aagaactcca aggacgccca gaaaatgatc aacgagatgc
2341 agaagcggaa ccggcagacc aacgagcgga tcgaggaaat catccggacc accggcaaag
2401 agaacgccaa gtacctgatc gagaagatca agctgcacga catgcaggaa ggcaagtgcc
2461 tgtacagcct ggaagccatc cctctggaag atctgctgaa caaccccttc aactatgagg
2521 tggaccacat catccccaga agcgtgtcct tcgacaacag cttcaacaac aaggtgctcg
2581 tgaagcagga agaagccagc aagaagggca accggacccc attccagtac ctgagcagca
2641 gcgacagcaa gatcagctac gaaaccttca agaagcacat cctgaatctg gccaagggca
2701 agggcagaat cagcaagacc aagaaagagt atctgctgga agaacgggac atcaacaggt
2761 tctccgtgca gaaagacttc atcaaccgga acctggtgga taccagatac gccaccagag
2821 gcctgatgaa cctgctgcgg agctacttca gagtgaacaa cctggacgtg aaagtgaagt
2881 ccatcaatgg cggcttcacc agctttctgc ggcggaagtg gaagtttaag aaagagcgga
2941 acaaggggta caagcaccac gccgaggacg ccctgatcat tgccaacgcc gatttcatct
3001 tcaaagagtg gaagaaactg gacaaggcca aaaaagtgat ggaaaaccag atgttcgagg
3061 aaaagcaggc cgagagcatg cccgagatcg aaaccgagca ggagtacaaa gagatcttca
3121 tcacccccca ccagatcaag cacattaagg acttcaagga ctacaagtac agccaccggg
3181 tggacaagaa gcctaataga gagctgatta acgacaccct gtactccacc cggaaggacg
3241 acaagggcaa caccctgatc gtgaacaatc tgaacggcct gtacgacaag gacaatgaca
3301 agctgaaaaa gctgatcaac aagagccccg aaaagctgct gatgtaccac cacgaccccc
3361 agacctacca gaaactgaag ctgattatgg aacagtacgg cgacgagaag aatcccctgt
3421 acaagtacta cgaggaaacc gggaactacc tgaccaagta ctccaaaaag gacaacggcc
3481 ccgtgatcaa gaagattaag tattacggca acaaactgaa cgcccatctg gacatcaccg
3541 acgactaccc caacagcaga aacaaggtcg tgaagctgtc cctgaagccc tacagattcg
3601 acgtgtacct ggacaatggc gtgtacaagt tcgtgaccgt gaagaatctg gatgtgatca
3661 aaaaagaaaa ctactacgaa gtgaatagca agtgctatga ggaagctaag aagctgaaga
3721 agatcagcaa ccaggccgag tttatcgcct ccttctacaa caacgatctg atcaagatca
3781 acggcgagct gtatagagtg atcggcgtga acaacgacct gctgaaccgg atcgaagtga
3841 acatgatcga catcacctac cgcgagtacc tggaaaacat gaacgacaag aggcccccca
3901 ggatcattaa gacaatcgcc tccaagaccc agagcattaa gaagtacagc acagacattc
3961 tgggcaacct gtatgaagtg aaatctaaga agcaccctca gatcatcaaa aagggcaaaa
4021 ggccggcggc cacgaaaaag gccggccagg caaaaaagaa aaagggatcc tacccatacg
4081 atgttccaga ttacgcttat ccctacgacg tgcctgatta tgcataccca tatgatgtcc
4141 ccgactatgc ctaagaattc ctagagctcg ctgatcagcc tcgactgtgc cttctagttg
4201 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
4261 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
4321 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag agaatagcag
4381 gcatgctggg gagctagagg ccgcaggaac ccctagtgat ggagttggcc actccctctc
4441 tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggcggcc
4501 tcagtgagcg agcgagcgcg cagctgcctg caggacatgt gagcaaaagg ccagcaaaag
4561 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
4621 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
4681 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
4741 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
4801 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
4861 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
4921 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
4981 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
5041 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
5101 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
5161 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
5221 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
5281 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
5341 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
5401 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
5461 ttaccatctg gccccagtgc tgcaatgata ccgcggcttc cacgctcacc ggctccagat
5521 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
5581 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
5641 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
5701 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
5761 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
5821 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
5881 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
5941 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
6001 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
6061 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
6121 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
6181 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
6241 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
6301 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
6361 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
6421 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
6481 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
6541 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
6601 aaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
6661 ttttttaacc aatagaccga aatcggcaaa atcccttata aatcaaaaga atagcccgag
6721 atagagttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
6781 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
6841 aaatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
6901 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
6961 gcgaaaggag cgggcgctaa ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
7021 acacccgccg cgcttaatgc gccgctacag ggcgcgtact atggttgctt tgacgtatgc
7081 ggtgtgaaat
//
