| 质粒名称: | pUC57-sgRNA |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 51132 |
| 存储实验室: | Xingxu Huang |
|
载体骨架基本信息 |
|
| 载体名称: | pUC57 |
| 载体出品公司: | -- |
| 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 |
| 筛选标记: | 新霉素（G418） |
| 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: | sgRNA scaffold |
| 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: | 限制性内切酶 |
| 5' 克隆位点: | AgeI(未破坏) |
| 3' 克隆位点: | NotI (未破坏) |
| 5' 测序引物 | CMV-F |
| 3' 测序引物 | SV40pA-R |
|
文献引用方法 |
|
| 材料和方法部分: | pUC57-sgRNA expression vector was a gift from Xingxu Huang (Addgene plasmid # 51132 ; http://n2t.net/addgene:51132 ; RRID:Addgene_51132) |
| 参考文献部分: | Efficient genome modification by CRISPR-Cas9 nickase with minimal off-target effects. Shen B, Zhang W, Zhang J, Zhou J, Wang J, Chen L, Wang L, Hodgkins A, Iyer V, Huang X, Skarnes WC. Nat Methods. 2014 Mar 2. doi: 10.1038/nmeth.2857. 10.1038/nmeth.2857 PubMed 24584192 |
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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 pUC57-sgRNA expression vector 2792 bp ds-DNA circular SYN 12-AUG-2019
DEFINITION For in vitro transcription of sgRNA from the T7 promoter..
ACCESSION .
VERSION .
KEYWORDS pUC57-sgRNA expression vector
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 2792)
AUTHORS Shen B, Zhang W, Zhang J, Zhou J, Wang J, Chen L, Wang L, Hodgkins
A, Iyer V, Huang X, Skarnes WC
TITLE Efficient genome modification by CRISPR-Cas9 nickase with minimal
off-target effects.
JOURNAL Nat Methods. 2014 Mar 2. doi: 10.1038/nmeth.2857.
PUBMED 24584192
REFERENCE 2 (bases 1 to 2792)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 12, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..2792
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 176..198
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 190..207
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 191..207
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
promoter 254..272
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
misc_RNA 294..369
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind complement(427..443)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(427..443)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(440..462)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 451..467
/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(475..505)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 520..541
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(659..676)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(830..1418)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(910..929)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(1589..2404)
/codon_start=1
/gene="aph(3')-Ia"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin in bacteria or G418
(Geneticin(R)) in eukaryotes"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
primer_bind 2312..2331
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
promoter complement(2405..2509)
/gene="bla"
/label=AmpR promoter
primer_bind 2577..2595
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind complement(2633..2655)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 2755..2774
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca
61 ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag
121 ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag
181 tcacgacgtt gtaaaacgac ggccagtgaa ttcgagctcg gtacctcgcg aatgcatcta
241 gatatcggat ccctaatacg actcactata ggtgagaccg agagagggtc tcagttttag
301 agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa agtggcaccg
361 agtcggtgct ttttttaaag ggcccgtcga ctgcagaggc ctgcatgcaa gcttggcgta
421 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
481 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
541 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
601 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgcggcc gccgcttcct
661 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
721 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
781 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
841 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
901 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
961 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
1021 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
1081 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
1141 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
1201 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
1261 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
1321 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
1381 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
1441 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
1501 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
1561 gtatatatga gtaaacttgg tctgacagtt agaaaaactc atcgagcatc aaatgaaact
1621 gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt ttctgtaatg
1681 aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat cggtctgcga
1741 ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa ataaggttat
1801 caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa agtttatgca
1861 tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa tcactcgcat
1921 caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt
1981 taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact gccagcgcat
2041 caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct gttttcccag
2101 ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc ttgatggtcg
2161 gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta acatcattgg
2221 caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc ccatacaatc
2281 gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac ccatataaat
2341 cagcatccat gttggaattt aatcgcggcc tagagcaaga cgtttcccgt tgaatatggc
2401 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
2461 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
2521 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
2581 ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac
2641 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
2701 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat
2761 cagagcagat tgtactgaga gtgcaccata tg
//
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