| 质粒名称: | pU6-Sp-pegRNA-HEK3_CTT_ins |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 132778 |
| 存储实验室: | David Liu |
|
载体骨架基本信息 |
|
| 载体名称: | pU6 |
| 载体抗性: | 氨苄青霉素 |
| 载体类型: | 哺乳细胞表达 |
| 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: | HEK3_CTT_ins pegRNA |
| 物种来源: | Synthetic |
| 插入大小: | 821 bp |
| 基因突变: | See manuscript |
| 启动子 | U6 |
| 克隆方法: | Ligation Independent克隆 |
| 5' 测序引物 | GACTATCATATGCTTACCGT |
|
文献引用方法 |
|
| 材料和方法部分: | pU6-Sp-pegRNA-HEK3_CTT_ins was a gift from David Liu (Addgene plasmid # 132778 ; http://n2t.net/addgene:132778 ; RRID:Addgene_132778) |
| 参考文献部分: | Search-and-replace genome editing without double-strand breaks or donor DNA. Anzalone AV, Randolph PB, Davis JR, Sousa AA, Koblan LW, Levy JM, Chen PJ, Wilson C, Newby GA, Raguram A, Liu DR. Nature. 2019 Oct 21. pii: 10.1038/s41586-019-1711-4. doi: 10.1038/s41586-019-1711-4. 10.1038/s41586-019-1711-4 PubMed 31634902 |
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载体基本信息
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载体简介
载体序列
LOCUS pU6-Sp-pegRNA-HEK3_CTT_ins 2305 bp ds-DNA circular SYN 18-OCT-2019
DEFINITION S. pyogenes pegRNA for CTT insertion at the HEK3 site of human cells
using prime editing.
ACCESSION .
VERSION .
KEYWORDS pU6-Sp-pegRNA-HEK3_CTT_ins
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 2305)
TITLE Liu Lab plasmids
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 2305)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Oct 18, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..2305
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_RNA 29..104
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
rep_origin complement(234..822)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(314..333)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(993..1853)
/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 1616..1635
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(1854..1958)
/gene="bla"
/label=AmpR promoter
promoter 2065..2305
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 2065..2085
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 2236..2255
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
ORIGIN
1 gaaacaccgg cccagactga gcacgtgagt tttagagcta gaaatagcaa gttaaaataa
61 ggctagtccg ttatcaactt gaaaaagtgg gaccgagtcg gtcctctgcc atcaaagcgt
121 gctcagtctg tttttttaag cttgggccgc tcgaggtacc tctctacata tgacatgtga
181 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
241 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
301 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
361 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
421 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
481 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
541 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
601 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
661 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
721 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
781 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
841 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
901 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
961 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
1021 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
1081 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
1141 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
1201 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
1261 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
1321 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
1381 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
1441 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
1501 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
1561 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
1621 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
1681 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
1741 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
1801 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
1861 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
1921 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
1981 cctgacgtcg ctagctgtac aaaaaagcag gctttaaagg aaccaattca gtcgactgga
2041 tccggtacca aggtcgggca ggaagagggc ctatttccca tgattccttc atatttgcat
2101 atacgataca aggctgttag agagataatt agaattaatt tgactgtaaa cacaaagata
2161 ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa
2221 ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg
2281 gctttatata tcttgtggaa aggac
//
