| 质粒名称: | ph7SK-gRNA |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 53189 |
| 存储实验室: | Charles Gersbach |
|
载体骨架基本信息 |
|
| 载体名称: | pzDonor |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | 3504 bp |
| 载体类型: | 哺乳细胞表达，CRISPR |
| 高拷贝/低拷贝: | 未知 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 启动子 |
human 7SK
|
| 克隆方法: | 限制性内切酶 |
| 3' 测序引物 | M13 Reverse |
|
文献引用方法 |
|
| 材料和方法部分: | ph7SK-gRNA was a gift from Charles Gersbach (爱迪基因 plasmid # 53189) |
| 参考文献部分: | Multiplex CRISPR/Cas9-based genome engineering from a single lentiviral vector. Kabadi AM, Ousterout DG, Hilton IB, Gersbach CA. Nucleic Acids Res. 2014 Aug 13. pii: gku749. 10.1093/nar/gku749 PubMed 25122746 |
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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 ph7SK-gRNA 3504 bp ds-DNA circular SYN 10-JAN-2018
DEFINITION Expresses the S. pyogenes sgRNA from the 7SK promoter.
ACCESSION .
VERSION .
KEYWORDS ph7SK-gRNA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 3504)
AUTHORS Kabadi AM, Ousterout DG, Hilton IB, Gersbach CA
TITLE Multiplex CRISPR/Cas9-based genome engineering from a single
lentiviral vector.
JOURNAL Nucleic Acids Res. 2014 Aug 13. pii: gku749.
PUBMED 25122746
COMMENT This file is created by Vector NTI
http://www.biofeng.com/
COMMENT ORIGDB|GenBank
COMMENT VNTAUTHORNAME|biofeng.com|
COMMENT VNTNAME|ph7SK-gRNA|
FEATURES Location/Qualifiers
source 1..3504
/organism="synthetic DNA construct"
/mol_type="other DNA"
rep_origin complement(3..458)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(140..161)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 352..371
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 585..607
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 599..616
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 600..616
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 626..645
/label=T7
/note="T7 promoter, forward primer"
promoter 626..644
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
misc_RNA 952..1027
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind complement(1083..1103)
/label=T3
/note="T3 promoter, forward primer"
promoter complement(1083..1101)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(1122..1138)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1122..1138)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(1135..1157)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 1146..1162
/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)."
promoter complement(1170..1200)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 1215..1236
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(1353..1370)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(1524..2112)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(1604..1623)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(2189..2983)
/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(2300..2319)
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
primer_bind 2910..2929
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
promoter complement(3018..3375)
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 3032..3167
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind complement(3086..3105)
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind 3332..3352
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter complement(3377..3481)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtacctt
661 taattctagt actatgcatc gtctcattgt ctgcagtatt tagcatgccc cacccatctg
721 caaggcattc tggatagtgt caaaacagcc ggaaatcaag tccgtttatc tcaaacttta
781 gcattttggg aataaatgat atttgctatg ctggttaaat tagattttag ttaaatttcc
841 tgctgaagct ctagtacgat aagcaacttg acctaagtgt aaagttgaga cttccttcag
901 gtttatatag cttgtgcgcc gcttgggtac ctcgggtctt cgagaagacc tgttttagag
961 ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag
1021 tcggtgcttt ttttctcgga ctgatgacct gagacgccgc ggtggagctc cagcttttgt
1081 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
1141 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
1201 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
1261 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
1321 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
1381 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
1441 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
1501 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
1561 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
1621 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
1681 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
1741 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
1801 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
1861 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
1921 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
1981 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
2041 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
2101 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
2161 aactcacgtt aagggatttt ggtcatgatc agaagaactc gtcaagaagg cgatagaagg
2221 cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt
2281 cgccgccaag ctcttcagca atatcacggg tagccaacgc tatgtcctga tagcggtccg
2341 ccacacccag ccggccacag tcgatgaatc cagaaaagcg gccattttcc accatgatat
2401 tcggcaagca ggcatcgcca tgggtcacga cgagatcctc gccgtcgggc atgctcgcct
2461 tgagcctggc gaacagttcg gctggcgcga gcccctgatg ctcttcgtcc agatcatcct
2521 gatcgacaag accggcttcc atccgagtac gtgctcgctc gatgcgatgt ttcgcttggt
2581 ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg ccgcattgca tcagccatga
2641 tggatacttt ctcggcagga gcaaggtgag atgacaggag atcctgcccc ggcacttcgc
2701 ccaatagcag ccagtccctt cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa
2761 cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc ttgcagttca ttcagggcac
2821 cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg
2881 cggcatcaga gcagccgatt gtctgttgtg cccagtcata gccgaatagc ctctccaccc
2941 aagcggccgg agaacctgcg tgcaatccat cttgttcaat catgcgaaac gatcctcatc
3001 ctgtctcttg atcgatcttt gcaaaagcct aggcctccaa aaaagcctcc tcactacttc
3061 tggaatagct cagaggccga ggcggcctcg gcctctgcat aaataaaaaa aattagtcag
3121 ccatggggcg gagaatgggc ggaactgggc ggagttaggg gcgggatggg cggagttagg
3181 ggcgggacta tggttgctga ctaattgaga tgcatgcttt gcatacttct gcctgctggg
3241 gagcctgggg actttccaca cctggttgct gactaattga gatgcatgct ttgcatactt
3301 ctgcctgctg gggagcctgg ggactttcca caccctaact gacacacatt ccacagctgg
3361 ttctttccgc ctcaggactc ttcctttttc aatattattg aagcatttat cagggttatt
3421 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
3481 gcacatttcc ccgaaaagtg ccac
//
