| 质粒名称: | pMA-SpCas9-g10 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 80793 |
| 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-g10 was a gift from Yonglun Luo (Addgene plasmid # 80793 ; http://n2t.net/addgene:80793 ; RRID:Addgene_80793) |
| 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-g10 2789 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION BsaI-based cloning of SpCas9 gRNA guide sequence, position 10/20/30
in the array.
ACCESSION .
VERSION .
KEYWORDS pMA-SpCas9-g10
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 278..300
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 292..309
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 293..309
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 320..339
/label=T7
/note="T7 promoter, forward primer"
promoter 320..338
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 403..643
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 403..423
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 574..593
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 670..745
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind complement(932..949)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(1042..1630)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(1122..1141)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(1801..2661)
/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 2424..2443
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(2662..2766)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
121 gatagggttg agtggccgct acagggcgct cccattcgcc attcaggctg cgcaactgtt
181 gggaagggcg tttcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
241 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
301 acggccagtg agcgcgacgt aatacgactc actatagggc gaattggcgg aaggccgtca
361 aggccacgtg tcttgtccag agctcggatc cggtctccgc tcgagggcct atttcccatg
421 attccttcat atttgcatat acgatacaag gctgttagag agataattgg aattaatttg
481 actgtaaaca caaagatatt agtacaaaat acgtgacgta gaaagtaata atttcttggg
541 tagtttgcag ttttaaaatt atgttttaaa atggactatc atatgcttac cgtaacttga
601 aagtatttcg atttcttggc tttatatatc ttgtggaaag gacgaaacac cgggtcttcg
661 agaagacctg ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact
721 tgaaaaagtg gcaccgagtc ggtgcttttt tgttttagag ctagaaatag caagttaggc
781 gggagacctg tacaacgcgt ggtacctgga gcacaagact ggcctcacgg gccttccgct
841 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaacat ggtcatagct
901 gtttccttgc gtattgggcg ctctccgctt cctcgctcac tgactcgctg cgctcggtcg
961 ttcgggtaaa gcctggggtg cctaatgagc aaaaggccag caaaaggcca ggaaccgtaa
1021 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
1081 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
1141 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
1201 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
1261 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
1321 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
1381 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
1441 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
1501 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
1561 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
1621 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
1681 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
1741 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
1801 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
1861 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
1921 cagtgctgca atgataccgc gagaaccacg ctcaccggct ccagatttat cagcaataaa
1981 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
2041 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
2101 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
2161 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
2221 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
2281 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
2341 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
2401 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
2461 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
2521 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
2581 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
2641 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
2701 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
2761 tccgcgcaca tttccccgaa aagtgccac
//
