| 质粒名称: | pMA-SpCas9-g3 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 80786 |
| 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-g3 was a gift from Yonglun Luo (Addgene plasmid # 80786 ; http://n2t.net/addgene:80786 ; RRID:Addgene_80786) |
| 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
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载体简介
载体序列
LOCUS pMA-SpCas9-g3 2789 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION BsaI-based cloning of SpCas9 gRNA guide sequence, position 3/13/23
in the array.
ACCESSION .
VERSION .
KEYWORDS pMA-SpCas9-g3
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"
primer_bind 60..77
/label=L4440
/note="L4440 vector, forward primer"
misc_RNA complement(264..339)
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter complement(366..606)
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind complement(416..435)
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
primer_bind complement(586..606)
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind complement(670..689)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(671..689)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(700..717)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(700..716)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(709..731)
/label=M13/pUC Forward
/note="In lacZ gene"
promoter 1032..1136
/gene="bla"
/label=AmpR promoter
CDS 1137..1997
/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(1355..1374)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 2168..2756
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 2657..2676
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 gccttttgct ggccttttgc tcattaggca ccccaggctt tacccgaacg accgagcgca
61 gcgagtcagt gagcgaggaa gcggagagcg cccaatacgc aaggaaacag ctatgaccat
121 gttaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cggaaggccc
181 gtgaggccag tcttgtgctc caggtaccac gcgttgtaca ggtctccctg gtaacttgct
241 atttctagct ctaaaacaaa aaagcaccga ctcggtgcca ctttttcaag ttgataacgg
301 actagcctta ttttaacttg ctatttctag ctctaaaaca ggtcttctcg aagacccggt
361 gtttcgtcct ttccacaaga tatataaagc caagaaatcg aaatactttc aagttacggt
421 aagcatatga tagtccattt taaaacataa ttttaaaact gcaaactacc caagaaatta
481 ttactttcta cgtcacgtat tttgtactaa tatctttgtg tttacagtca aattaattcc
541 aattatctct ctaacagcct tgtatcgtat atgcaaatat gaaggaatca tgggaaatag
601 gccctcgtcc tgagaccgga tccgagctct ggacaagaca cgtggccttg acggccttcc
661 gccaattcgc cctatagtga gtcgtattac gtcgcgctca ctggccgtcg ttttacaacg
721 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt
781 cgccagctgg cgtaatagcg aagaggcccg caccgaaacg cccttcccaa cagttgcgca
841 gcctgaatgg cgaatgggag cgccctgtag cggccactca accctatctc ggtctattct
901 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
961 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
1021 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
1081 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
1141 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
1201 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
1261 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
1321 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
1381 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
1441 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
1501 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
1561 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
1621 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
1681 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
1741 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
1801 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt ggttctcgcg
1861 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
1921 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
1981 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
2041 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
2101 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
2161 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
2221 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
2281 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
2341 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
2401 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
2461 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
2521 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
2581 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
2641 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
2701 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
2761 ccagcaacgc ggccttttta cggttcctg
//
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