| 质粒名称: | gRNA_AAVS1-T2 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 41818 |
| 存储实验室: | George Church |
|
载体骨架基本信息 |
|
| 载体名称: | pCR-Blunt II-TOPO |
| 载体出品公司: | Invitrogen |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 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: | -- |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | gRNA_AAVS1-T2 |
| 物种来源: | -- |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | TOPO克隆 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | T7 |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | gRNA_AAVS1-T2 was a gift from George Church (Addgene plasmid # 41818 ; http://n2t.net/addgene:41818 ; RRID:Addgene_41818) |
| 参考文献部分: | RNA-Guided Human Genome Engineering via Cas9. Mali P, Yang L, Esvelt KM, Aach J, Guell M, Dicarlo JE, Norville JE, Church GM. Science. 2013 Jan 3. 10.1126/science.1232033 PubMed 23287722 |
买家导航
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 gRNA_AAVS1-T2 3973 bp ds-DNA circular SYN 07-APR-2020
DEFINITION Expresses a guide RNA (gRNA) to target human AAVS1 (T2 target
sequence) for genome engineering.
ACCESSION .
VERSION .
KEYWORDS gRNA_AAVS1-T2
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 3973)
AUTHORS Mali P, Yang L, Esvelt KM, Aach J, Guell M, Dicarlo JE, Norville JE,
Church GM
TITLE RNA-Guided Human Genome Engineering via Cas9.
JOURNAL Science. 2013 Jan 3.
PUBMED 23287722
REFERENCE 2 (bases 1 to 3973)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 7, 2020 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..3973
/organism="synthetic DNA construct"
/mol_type="other DNA"
protein_bind 107..128
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 143..173
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 181..197
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 186..208
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 205..221
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 205..221
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
promoter 239..257
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 239..256
/label=SP6
/note="SP6 promoter, forward primer"
protein_bind 342..366
/gene="mutant version of attB"
/label=attB2
/bound_moiety="BP Clonase(TM)"
/note="recombination site for the Gateway(R) BP reaction"
misc_RNA complement(378..453)
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter complement(482..722)
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind complement(532..551)
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
primer_bind complement(702..722)
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind complement(861..880)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(862..880)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(887..904)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(887..903)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(896..918)
/label=M13/pUC Forward
/note="In lacZ gene"
CDS 1041..1343
/codon_start=1
/gene="ccdB"
/product="CcdB, a bacterial toxin that poisons DNA gyrase"
/label=ccdB
/note="Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains."
/translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
primer_bind 1222..1241
/label=ccdB-fwd
/note="ccdB gene, forward primer"
CDS 1692..2486
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(1746..1765)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 2356..2375
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
regulatory 2686..2695
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2692..3066
/codon_start=1
/gene="Sh ble from Streptoalloteichus hindustanus"
/product="antibiotic-binding protein"
/label=BleoR
/note="confers resistance to bleomycin, phleomycin, and
Zeocin(TM)"
/translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
EFALRDPAGNCVHFVAEEQD"
rep_origin 3204..3792
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3693..3712
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 3946..3963
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctat
241 ttaggtgaca ctatagaata ctcaagctat gcatcaagct tggtaccgag ctcggatcca
301 ctagtaacgg ccgccagtgt gctggaattc gccctttaat gccaactttg tacaagaaag
361 ctgggtctag aaaaaaagca ccgactcggt gccacttttt caagttgata acggactagc
421 cttattttaa cttgctattt ctagctctaa aacatcctgt ccctagtggc cccggtgttt
481 cgtcctttcc acaagatata taaagccaag aaatcgaaat actttcaagt tacggtaagc
541 atatgatagt ccattttaaa acataatttt aaaactgcaa actacccaag aaattattac
601 tttctacgtc acgtattttg tactaatatc tttgtgttta cagtcaaatt aattctaatt
661 atctctctaa cagccttgta tcgtatatgc aaatatgaag gaatcatggg aaataggccc
721 tcttcctgcc cgaccttggt accggatcca gtcgactgaa ttggttcctt taaagcctgc
781 ttttttgtac aaagggcgaa ttctgcagat atccatcaca ctggcggccg ctcgagcatg
841 catctagagg gcccaattcg ccctatagtg agtcgtatta caattcactg gccgtcgttt
901 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
961 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
1021 tgcgcagcct atacgtacgg cagtttaagg tttacaccta taaaagagag agccgttatc
1081 gtctgtttgt ggatgtacag agtgatatta ttgacacgcc ggggcgacgg atggtgatcc
1141 ccctggccag tgcacgtctg ctgtcagata aagtctcccg tgaactttac ccggtggtgc
1201 atatcgggga tgaaagctgg cgcatgatga ccaccgatat ggccagtgtg ccggtctccg
1261 ttatcgggga agaagtggct gatctcagcc accgcgaaaa tgacatcaaa aacgccatta
1321 acctgatgtt ctggggaata taaatgtcag gcatgagatt atcaaaaagg atcttcacct
1381 agatcctttt cacgtagaaa gccagtccgc agaaacggtg ctgaccccgg atgaatgtca
1441 gctactgggc tatctggaca agggaaaacg caagcgcaaa gagaaagcag gtagcttgca
1501 gtgggcttac atggcgatag ctagactggg cggttttatg gacagcaagc gaaccggaat
1561 tgccagctgg ggcgccctct ggtaaggttg ggaagccctg caaagtaaac tggatggctt
1621 tcttgccgcc aaggatctga tggcgcaggg gatcaagctc tgatcaagag acaggatgag
1681 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
1741 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
1801 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
1861 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
1921 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
1981 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
2041 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
2101 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
2161 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
2221 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
2281 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
2341 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
2401 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
2461 atcgccttct tgacgagttc ttctgaatta ttaacgctta caatttcctg atgcggtatt
2521 ttctccttac gcatctgtgc ggtatttcac accgcatcag gtggcacttt tcggggaaat
2581 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
2641 agacaataac cctgataaat gcttcaataa tagcacgtga ggagggccac catggccaag
2701 ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg
2761 accgaccggc tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg
2821 gacgacgtga ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg
2881 gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc
2941 acgaacttcc gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg
3001 cgggagttcg ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag
3061 gactgacacg tgctaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
3121 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
3181 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
3241 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
3301 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
3361 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
3421 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
3481 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
3541 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
3601 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
3661 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
3721 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
3781 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
3841 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
3901 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
3961 gaggaagcgg aag
//
