| 质粒名称: | pMA-SpCas9-g4 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 80787 |
| 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-g4 was a gift from Yonglun Luo (Addgene plasmid # 80787 ; http://n2t.net/addgene:80787 ; RRID:Addgene_80787) |
| 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-g4 2789 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION BsaI-based cloning of SpCas9 gRNA guide sequence, position 4/14/24
in the array.
ACCESSION .
VERSION .
KEYWORDS pMA-SpCas9-g4
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"
CDS complement(607..1467)
/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 1230..1249
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(1468..1572)
/gene="bla"
/label=AmpR promoter
primer_bind 1873..1895
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 1887..1904
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 1888..1904
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 1915..1934
/label=T7
/note="T7 promoter, forward primer"
promoter 1915..1933
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 1998..2238
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 1998..2018
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 2169..2188
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 2265..2340
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind complement(2527..2544)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2637..436)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2717..2736)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
61 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
121 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
181 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
241 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
301 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
361 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
421 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
481 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
541 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
601 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
661 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
721 tggccccagt gctgcaatga taccgcgaga accacgctca ccggctccag atttatcagc
781 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
841 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
901 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
961 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
1021 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
1081 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
1141 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
1201 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
1261 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
1321 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
1381 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
1441 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
1501 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
1561 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
1621 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
1681 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgg ccgctacagg
1741 gcgctcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgtttcg gtgcgggcct
1801 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
1861 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgagcgc gacgtaatac
1921 gactcactat agggcgaatt ggcggaaggc cgtcaaggcc acgtgtcttg tccagagctc
1981 ggatccggtc tcgccaggag ggcctatttc ccatgattcc ttcatatttg catatacgat
2041 acaaggctgt tagagagata attggaatta atttgactgt aaacacaaag atattagtac
2101 aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta aaattatgtt
2161 ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc ttggctttat
2221 atatcttgtg gaaaggacga aacaccgggt cttcgagaag acctgtttta gagctagaaa
2281 tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc gagtcggtgc
2341 ttttttgttt tagagctaga aatagcaagt tatgttggag acctgtacaa cgcgtggtac
2401 ctggagcaca agactggcct cacgggcctt ccgctcactg cccgctttcc agtcgggaaa
2461 cctgtcgtgc cagctgcatt aacatggtca tagctgtttc cttgcgtatt gggcgctctc
2521 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg gtaaagcctg gggtgcctaa
2581 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
2641 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
2701 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
2761 cctgttccga ccctgccgct taccggata
//
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