| 质粒名称: | pMA-SpCas9-g5 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 80788 |
| 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-g5 was a gift from Yonglun Luo (Addgene plasmid # 80788 ; http://n2t.net/addgene:80788 ; RRID:Addgene_80788) |
| 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-g5 2789 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION BsaI-based cloning of SpCas9 gRNA guide sequence, position 5/15/25
in the array.
ACCESSION .
VERSION .
KEYWORDS pMA-SpCas9-g5
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 39..56
/label=L4440
/note="L4440 vector, forward primer"
misc_RNA complement(243..318)
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter complement(345..585)
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind complement(395..414)
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
primer_bind complement(565..585)
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind complement(649..668)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(650..668)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(679..696)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(679..695)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(688..710)
/label=M13/pUC Forward
/note="In lacZ gene"
promoter 1011..1115
/gene="bla"
/label=AmpR promoter
CDS 1116..1976
/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(1334..1353)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 2147..2735
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 2636..2655
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 cattaggcac cccaggcttt acccgaacga ccgagcgcag cgagtcagtg agcgaggaag
61 cggagagcgc ccaatacgca aggaaacagc tatgaccatg ttaatgcagc tggcacgaca
121 ggtttcccga ctggaaagcg ggcagtgagc ggaaggcccg tgaggccagt cttgtgctcc
181 aggtaccacg cgttgtacag gtctcctgca taacttgcta tttctagctc taaaacaaaa
241 aagcaccgac tcggtgccac tttttcaagt tgataacgga ctagccttat tttaacttgc
301 tatttctagc tctaaaacag gtcttctcga agacccggtg tttcgtcctt tccacaagat
361 atataaagcc aagaaatcga aatactttca agttacggta agcatatgat agtccatttt
421 aaaacataat tttaaaactg caaactaccc aagaaattat tactttctac gtcacgtatt
481 ttgtactaat atctttgtgt ttacagtcaa attaattcca attatctctc taacagcctt
541 gtatcgtata tgcaaatatg aaggaatcat gggaaatagg ccctcaacag gagaccggat
601 ccgagctctg gacaagacac gtggccttga cggccttccg ccaattcgcc ctatagtgag
661 tcgtattacg tcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
721 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
781 agaggcccgc accgaaacgc ccttcccaac agttgcgcag cctgaatggc gaatgggagc
841 gccctgtagc ggccactcaa ccctatctcg gtctattctt ttgatttata agggattttg
901 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
961 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
1021 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
1081 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
1141 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
1201 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
1261 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
1321 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
1381 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
1441 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
1501 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
1561 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
1621 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
1681 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
1741 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
1801 ttgctgataa atctggagcc ggtgagcgtg gttctcgcgg tatcattgca gcactggggc
1861 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
1921 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
1981 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
2041 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
2101 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
2161 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
2221 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
2281 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
2341 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
2401 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
2461 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
2521 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
2581 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
2641 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
2701 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
2761 ggttcctggc cttttgctgg ccttttgct
//
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