| 质粒名称: | pMA-SpCas9-g1 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 80784 |
| 存储实验室: | Yonglun Luo |
|
载体骨架基本信息 |
|
| 载体名称: | pMA15 |
| 载体出品公司: | Lifetechnologies |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 2380 bp |
| 完整质粒大小: | 2789 bp |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达,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: | CRISPR gRNA expression cassette (for SpCas9) |
| 物种来源: | Synthetic |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | U6 |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | BamHI(未破坏) |
| 3' 克隆位点: | BsrGI (未破坏) |
| 5' 测序引物 | TAATACGACTCACTATAGG |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pMA-SpCas9-g1 was a gift from Yonglun Luo (Addgene plasmid # 80784 ; http://n2t.net/addgene:80784 ; RRID:Addgene_80784) |
| 参考文献部分: | Golden Gate Assembly of CRISPR gRNA expression array for simultaneously targeting multiple genes. Vad-Nielsen J, Lin L, Bolund L, Nielsen AL, Luo Y. Cell Mol Life Sci. 2016 May 13. 10.1007/s00018-016-2271-5 [pii] PubMed 27178736 |
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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 pMA-SpCas9-g1 2789 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION BsaI-based cloning of SpCas9 gRNA guide sequence, position 1/11/21
in the array.
ACCESSION .
VERSION .
KEYWORDS pMA-SpCas9-g1
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 2789)
AUTHORS Vad-Nielsen J, Lin L, Bolund L, Nielsen AL, Luo Y
TITLE Golden Gate Assembly of CRISPR gRNA expression array for
simultaneously targeting multiple genes.
JOURNAL Cell Mol Life Sci. 2016 May 13.
PUBMED 27178736
REFERENCE 2 (bases 1 to 2789)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..2789
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 42..282
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 42..62
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 213..232
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 309..384
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind complement(571..588)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(681..1269)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(761..780)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(1440..2300)
/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 2063..2082
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(2301..2405)
/gene="bla"
/label=AmpR promoter
primer_bind 2706..2728
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 2720..2737
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 2721..2737
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 2748..2767
/label=T7
/note="T7 promoter, forward primer"
promoter 2748..2766
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
1 ggccacgtgt cttgtccaga gctcggatcc ggtctcgcta tgagggccta tttcccatga
61 ttccttcata tttgcatata cgatacaagg ctgttagaga gataattgga attaatttga
121 ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt
181 agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa
241 agtatttcga tttcttggct ttatatatct tgtggaaagg acgaaacacc gggtcttcga
301 gaagacctgt tttagagcta gaaatagcaa gttaaaataa ggctagtccg ttatcaactt
361 gaaaaagtgg caccgagtcg gtgctttttt gttttagagc tagaaatagc aagttacatg
421 ggagacctgt acaacgcgtg gtacctggag cacaagactg gcctcacggg ccttccgctc
481 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaacatg gtcatagctg
541 tttccttgcg tattgggcgc tctccgcttc ctcgctcact gactcgctgc gctcggtcgt
601 tcgggtaaag cctggggtgc ctaatgagca aaaggccagc aaaaggccag gaaccgtaaa
661 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
721 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
781 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
841 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
901 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
961 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
1021 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
1081 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
1141 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
1201 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
1261 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
1321 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
1381 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
1441 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
1501 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
1561 agtgctgcaa tgataccgcg agaaccacgc tcaccggctc cagatttatc agcaataaac
1621 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
1681 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
1741 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
1801 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
1861 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
1921 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
1981 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
2041 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
2101 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
2161 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
2221 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
2281 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
2341 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
2401 ccgcgcacat ttccccgaaa agtgccacct aaattgtaag cgttaatatt ttgttaaaat
2461 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa
2521 tcccttataa atcaaaagaa tagaccgaga tagggttgag tggccgctac agggcgctcc
2581 cattcgccat tcaggctgcg caactgttgg gaagggcgtt tcggtgcggg cctcttcgct
2641 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
2701 gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgacgtaa tacgactcac
2761 tatagggcga attggcggaa ggccgtcaa
//
