| 质粒名称: | PX552 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 60958 |
| 存储实验室: | Feng Zhang |
|
载体骨架基本信息 |
|
| 载体名称: | pAAV |
| 载体抗性: | 氨苄霉素 |
| 空载体大小: | 2905 bp |
| 完整质粒大小: | 5314 bp |
| 载体类型: | 哺乳细胞表达，Mouse Targeting, AAV, 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: | U6_(SpaI)_sgRNA |
| 物种来源: | Synthetic |
| 插入大小: | 351 bp |
| 启动子 | U6 |
| 克隆方法: | Gibson Cloning |
| 5' 测序引物 | cgcgtgagggcctatttcc |
| 3' 测序引物 | gcaagtgggttttaggaccag |
|
文献引用方法 |
|
| 材料和方法部分: | PX552 was a gift from Feng Zhang (爱迪基因 plasmid # 60958) |
| 参考文献部分: | n vivo interrogation of gene function in the mammalian brain using CRISPR-Cas9. Swiech L, Heidenreich M, Banerjee A, Habib N, Li Y, Trombetta J, Sur M, Zhang F. Nat Biotechnol. 2014 Oct 19. doi: 10.1038/nbt.3055. 10.1038/nbt.3055 PubMed 25326897 |
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 PX552 5303 bp ds-DNA circular SYN 10-JAN-2018
DEFINITION pAAV-U6sgRNA(SapI)_hSyn-GFP-KASH-bGH (SpGuide acceptor). AAV plasmid
for sgRNA cloning. GFP-KASH fusion facilitates FACS sorting of cells
and nuclei. .
ACCESSION .
VERSION .
KEYWORDS PX552
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5303)
AUTHORS Swiech L, Heidenreich M, Banerjee A, Habib N, Li Y, Trombetta J, Sur
M, Zhang F
TITLE In vivo interrogation of gene function in the mammalian brain using
CRISPR-Cas9.
JOURNAL Nat Biotechnol. 2014 Oct 19. doi: 10.1038/nbt.3055.
PUBMED 25326897
COMMENT This file is created by Vector NTI
http://www.biofeng.com/
COMMENT ORIGDB|GenBank
COMMENT VNTAUTHORNAME|biofeng.com|
COMMENT VNTNAME|PX552|
FEATURES Location/Qualifiers
source 1..5303
/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 423..498
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter 532..979
/label=hSyn promoter
/note="human synapsin I promoter; confers neuron-specific
expression (Kugler et al., 2003)"
regulatory 1011..1020
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 1017..1733
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
primer_bind complement(1062..1083)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(1323..1342)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 1670..1691
/label=EGFP-C
/note="EGFP, forward primer"
polyA_signal 2050..2526
/label=hGH poly(A) signal
/note="human growth hormone polyadenylation signal"
primer_bind complement(2249..2268)
/label=hGH-PA-R
/note="Human growth hormone terminator, reverse primer"
repeat_region 2566..2706
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 2566..2695
/label=AAV2 ITR
rep_origin 2781..3236
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2868..2887)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 3078..3099
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(3253..3272)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 3372..3394
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(3432..3450)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 3518..3622
/gene="bla"
/label=AmpR promoter
CDS 3623..4483
/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(3841..3860)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 4654..5242
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 5143..5162
/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 caccggaaga gcgagctctt
421 ctgttttaga gctagaaata gcaagttaaa ataaggctag tccgttatca acttgaaaaa
481 gtggcaccga gtcggtgctt tttttctaga ctgcagaggg ccctgcgtat gagtgcaagt
541 gggttttagg accaggatga ggcggggtgg gggtgcctac ctgacgaccg accccgaccc
601 actggacaag cacccaaccc ccattcccca aattgcgcat cccctatcag agagggggag
661 gggaaacagg atgcggcgag gcgcgtgcgc actgccagct tcagcaccgc ggacagtgcc
721 ttcgcccccg cctggcggcg cgcgccaccg ccgcctcagc actgaaggcg cgctgacgtc
781 actcgccggt cccccgcaaa ctccccttcc cggccacctt ggtcgcgtcc gcgccgccgc
841 cggcccagcc ggaccgcacc acgcgaggcg cgagataggg gggcacgggc gcgaccatct
901 gcgctgcggc gccggcgact cagcgctgcc tcagtctgcg gtgggcagcg gaggagtcgt
961 gtcgtgcctg agagcgcagt cgagaaggta ccggatcctc tagagtcgac gccaccatgg
1021 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
1081 acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca
1141 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
1201 tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc
1261 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca
1321 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
1381 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
1441 tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca
1501 tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc
1561 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
1621 tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
1681 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtccggac
1741 tcagatctcg agaggaggag gaggagacag acagcaggat gccccacctc gacagccccg
1801 gcagctccca gccgagacgc tccttcctct caagggtgat cagggcagcg ctaccgttgc
1861 agctgcttct gctgctgctg ctgctcctgg cctgcctgct acctgcctct gaagatgact
1921 acagctgcac ccaggccaac aactttgccc gatccttcta ccccatgctg cggtacacca
1981 acgggccacc tcccacctag gaattcgata tcaagcttat cgataccgag cgctgctcga
2041 gagatctacg ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc cctggaagtt
2101 gccactccag tgcccaccag ccttgtccta ataaaattaa gttgcatcat tttgtctgac
2161 taggtgtcct tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcaagtt
2221 gggaagacaa cctgtagggc ctgcggggtc tattgggaac caagctggag tgcagtggca
2281 caatcttggc tcactgcaat ctccgcctcc tgggttcaag cgattctcct gcctcagcct
2341 cccgagttgt tgggattcca ggcatgcatg accaggctca gctaattttt gtttttttgg
2401 tagagacggg gtttcaccat attggccagg ctggtctcca actcctaatc tcaggtgatc
2461 tacccacctt ggcctcccaa attgctggga ttacaggcgt gaaccactgc tcccttccct
2521 gtccttctga ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga
2581 tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg
2641 tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc
2701 tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
2761 tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
2821 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
2881 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
2941 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg
3001 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
3061 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
3121 gggctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
3181 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg
3241 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc
3301 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
3361 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
3421 gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
3481 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
3541 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
3601 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
3661 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
3721 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
3781 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
3841 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
3901 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
3961 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
4021 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
4081 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
4141 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
4201 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
4261 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
4321 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
4381 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
4441 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
4501 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
4561 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
4621 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
4681 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
4741 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
4801 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
4861 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
4921 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
4981 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
5041 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
5101 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
5161 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
5221 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
5281 ttgctggcct tttgctcaca tgt
//
