| 质粒名称: | AAV:ITR-U6-sgRNA(LacZ)-pCBh-Cre-WPRE-hGHpA-ITR |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 60228 |
| 存储实验室: | Feng Zhang |
|
载体骨架基本信息 |
|
| 载体名称: | AAV |
| 载体抗性: | 氨苄西林 |
| 空载体大小: | 2597 bp |
| 完整质粒大小: | 6397 bp |
| 载体类型: | 哺乳动物表达，小鼠靶向，AAV，CRE/LOX，CRISPR |
| 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: |
sgRNA
|
| 备用名称： | single guide RNA、guide RNA、gRNA、 |
| 物种来源: | 光合细菌 Rhodopseudomonas palustris |
| 插入大小: | 102 bp |
| 启动子 | U6 |
| 克隆方法: | 限制性酶 |
| 5' 克隆位点: | Mlui（not destroyed） |
| 3' 克隆位点: | Xbai（未销毁） |
| 5' 测序引物 | GAGGGCCTATTTCCCATGATTC |
| 3' 测序引物 | NA |
|
文献引用方法 |
|
| 材料和方法部分: | AAV:ITR-U6-sgRNA(LacZ)-pCBh-Cre-WPRE-hGHpA-ITR was a gift from Feng Zhang (Addgene plasmid # 60228 ; http://n2t.net/addgene:60228 ; RRID:Addgene_60228) |
| 参考文献部分: | CRISPR-Cas9 Knockin Mice for Genome Editing and Cancer Modeling. Platt RJ, Chen S, Zhou Y, Yim MJ, Swiech L, Kempton HR, Dahlman JE, Parnas O, Eisenhaure TM, Jovanovic M, Graham DB, Jhunjhunwala S, Heidenreich M, Xavier RJ, Langer R, Anderson DG, Hacohen N, Regev A, Feng G, Sharp PA, Zhang F. Cell. 2014 Sep 24. pii: S0092-8674(14)01163-5. doi: 10.1016/j.cell.2014.09.014. 10.1016/j.cell.2014.09.014 PubMed 25263330 |
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
-pCBh-Cre-WPRE-hGHpA-ITR质粒图谱.png)
载体简介
载体序列
LOCUS AAV:ITR-U6-sgRNA(LacZ)-pCBh-Cre-WPRE-hGHpA-ITR 6379 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Expresses Cre recombinase from the Cbh promoter and one U6-driven
sgRNA control targeting LacZ. AAV backbone..
ACCESSION .
VERSION .
KEYWORDS AAV:ITR-U6-sgRNA(LacZ)-pCBh-Cre-WPRE-hGHpA-ITR
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6379)
AUTHORS Platt RJ, Chen S, Zhou Y, Yim MJ, Swiech L, Kempton HR, Dahlman JE,
Parnas O, Eisenhaure TM, Jovanovic M, Graham DB, Jhunjhunwala S,
Heidenreich M, Xavier RJ, Langer R, Anderson DG, Hacohen N, Regev A,
Feng G, Sharp PA, Zhang F
TITLE CRISPR-Cas9 Knockin Mice for Genome Editing and Cancer Modeling.
JOURNAL Cell. 2014 Sep 24. pii: S0092-8674(14)01163-5. doi:
10.1016/j.cell.2014.09.014.
PUBMED 25263330
REFERENCE 2 (bases 1 to 6379)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6379
/organism="synthetic DNA construct"
/mol_type="other DNA"
repeat_region 1..141
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 12..141
/label=AAV2 ITR
promoter 156..396
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 156..176
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 327..346
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 426..501
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
enhancer 591..876
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer;
contains an 18-bp deletion relative to the standard CMV
enhancer"
promoter 878..1155
/label=chicken beta-actin promoter
intron 1156..1384
/label=hybrid intron
/note="hybrid between chicken beta-actin (CBA) and minute
virus of mice (MMV) introns (Gray et al., 2011)"
CDS 1405..1425
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 2452..2478
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
misc_feature 2506..3094
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(2559..2579)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(2977..2988)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
polyA_signal 3126..3602
/label=hGH poly(A) signal
/note="human growth hormone polyadenylation signal"
primer_bind complement(3325..3344)
/label=hGH-PA-R
/note="Human growth hormone terminator, reverse primer"
repeat_region 3642..3782
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 3642..3771
/label=AAV2 ITR
rep_origin 3857..4312
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(3944..3963)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4154..4175
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(4329..4348)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 4448..4470
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(4508..4526)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 4594..4698
/gene="bla"
/label=AmpR promoter
CDS 4699..5559
/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 complement(4917..4936)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 5730..6318
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6219..6238
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
121 actccatcac taggggttcc tgcggccgca cgcgtgaggg cctatttccc atgattcctt
181 catatttgca tatacgatac aaggctgtta gagagataat tggaattaat ttgactgtaa
241 acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg
301 cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt
361 tcgatttctt ggctttatat atcttgtgga aaggacgaaa caccgtgcga atacgcccac
421 gcgatgtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa
481 aaagtggcac cgagtcggtg cttttttgtt ttagagctag aaatagcaag ttaaaataag
541 gctagtccgt agcgcgtgcg ccaattctgc agacaaattc tagaggtacc cgttacataa
601 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
661 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
721 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
781 ggtaaatggc ccgcctggca ttgtgcccag tacatgacct tatgggactt tcctacttgg
841 cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc cacgttctgc
901 ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt tattttttaa
961 ttattttgtg cagcgatggg ggcggggggg gggggggggc gcgcgccagg cggggcgggg
1021 cggggcgagg ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc
1081 gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga
1141 agcgcgcggc gggcgggagt cgctgcgacg ctgccttcgc cccgtgcccc gctccgccgc
1201 cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg tgagcgggcg
1261 ggacggccct tctcctccgg gctgtaatta gctgagcaag aggtaagggt ttaagggatg
1321 gttggttggt ggggtattaa tgtttaatta cctggagcac ctgcctgaaa tcactttttt
1381 tcaggttgga ccggtgccac catgcccaag aagaagagga aggtgtccaa tctcctgact
1441 gttcaccaga acctccctgc gctgccagta gatgccacta gcgatgaggt caggaaaaat
1501 ctcatggata tgtttaggga tagacaggcg ttttctgaac acacctggaa aatgctgctt
1561 agcgtgtgcc gatcctgggc agcctggtgt aagctgaaca atcgcaaatg gttccccgcc
1621 gagccggagg acgtgcgcga ttacctgctg tatctccagg caagagggct ggctgtcaag
1681 actatccagc agcacttggg ccaactgaat atgctgcatc gacgcagcgg gctcccccgg
1741 cctagcgatt caaacgcagt ctcccttgtt atgaggagaa ttagaaagga aaacgtagat
1801 gcgggtgaga gggctaagca ggctctcgct tttgagcgga ctgatttcga ccaggtcaga
1861 tccctgatgg agaacagcga tcggtgccag gacatcagga acctcgcatt tctgggaatt
1921 gcatataaca cacttctgcg catagctgag atcgcccgga tcagagtgaa agacatcagt
1981 cgaacggacg gcggccggat gcttattcat attggacgca caaagacatt ggtcagcacc
2041 gctggcgttg aaaaggcctt gtccctgggc gtaacgaagc tggtggaaag atggatctca
2101 gtgtccggcg tggctgacga ccctaataat tacttgttct gtcgagtgag aaaaaacgga
2161 gtcgccgcgc cctctgccac cagccaattg agtacacggg cccttgaagg gatctttgag
2221 gcaacccacc gactcatata cggagccaag gatgacagtg gccagaggta tctcgcctgg
2281 tcaggtcatt ctgctagggt gggggccgca cgagacatgg cgcgggcagg agtctccata
2341 ccagagatta tgcaagctgg aggttggaca aatgtgaaca tcgttatgaa ctatatccgc
2401 aatcttgact ctgaaaccgg ggccatggtg agactgctcg aagatggtga ctacccatac
2461 gatgttccag attacgctta agaattcgat atcaagctta tcgataatca acctctggat
2521 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
2581 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
2641 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
2701 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
2761 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
2821 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
2881 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta tgttgccacc
2941 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
3001 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
3061 acgagtcgga tctccctttg ggccgcctcc ccgcatcgat accgagcgct gctcgagaga
3121 tctacgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg gaagttgcca
3181 ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg tctgactagg
3241 tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg caagttggga
3301 agacaacctg tagggcctgc ggggtctatt gggaaccaag ctggagtgca gtggcacaat
3361 cttggctcac tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct cagcctcccg
3421 agttgttggg attccaggca tgcatgacca ggctcagcta atttttgttt ttttggtaga
3481 gacggggttt caccatattg gccaggctgg tctccaactc ctaatctcag gtgatctacc
3541 caccttggcc tcccaaattg ctgggattac aggcgtgaac cactgctccc ttccctgtcc
3601 ttctgatttt gtaggtaacc acgtgcggac cgagcggccg caggaacccc tagtgatgga
3661 gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc
3721 ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca
3781 ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
3841 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
3901 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
3961 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
4021 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat
4081 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
4141 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc
4201 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
4261 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc
4321 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
4381 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
4441 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
4501 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
4561 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
4621 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
4681 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
4741 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
4801 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
4861 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
4921 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
4981 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
5041 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
5101 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
5161 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
5221 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
5281 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
5341 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
5401 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
5461 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
5521 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
5581 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
5641 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
5701 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
5761 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
5821 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
5881 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
5941 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
6001 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
6061 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
6121 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
6181 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
6241 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
6301 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
6361 tggccttttg ctcacatgt
//
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