Abstract：
:CRISPR/Cas9 is a powerful genome editing system but uncontrolled Cas9 nuclease expression triggers off-target effects and even in vivo immune responses. Inspired by synthetic biology, here we built a synthetic switch that self-regulates Cas9 expression not only in the transcription step by guide RNA-aided self-cleavage of cas9 gene, but also in the translation step by L7Ae:K-turn repression system. We showed that the synthetic switch enabled simultaneous transcriptional and translational repression, hence stringently attenuating the Cas9 expression. The restricted Cas9 expression induced high efficiency on-target indel mutation while minimizing the off-target effects. Furthermore, we unveiled the correlation between Cas9 expression kinetics and on-target/off-target mutagenesis. The synthetic switch conferred detectable Cas9 expression and concomitant high frequency on-target mutagenesis at as early as 6 h, and restricted the Cas9 expression and off-target effects to minimal levels through 72 h. The synthetic switch is compact enough to be incorporated into viral vectors for self-regulation of Cas9 expression, thereby providing a novel 'hit and run' strategy for in vivo genome editing.
journal_name
Nucleic Acids Resjournal_title
Nucleic acids researchauthors
Shen CC,Hsu MN,Chang CW,Lin MW,Hwu JR,Tu Y,Hu YCdoi
10.1093/nar/gky1165subject
Has Abstractpub_date
2019-02-20 00:00:00pages
e13issue
3eissn
0305-1048issn
1362-4962pii
5193335journal_volume
47pub_type
杂志文章abstract：:Partial restriction digestion is used to map restriction sites and the location of genes within yeast artificial chromosomes (YACs). Locus-specific probes are hybridised to the partially digested YAC DNA and the fragments to which they hybridise are compared with the pattern of partial digestion products that include ...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/24.22.4495
更新日期：1996-11-15 00:00:00
abstract：:LOCtarget is a web server and database that predicts and annotates sub-cellular localization for structural genomics targets; LOCnet is one of the methods used in LOCtarget that can predict sub-cellular localization for all eukaryotic and prokaryotic proteins. Targets are taken from the central registration database f...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkh441
更新日期：2004-07-01 00:00:00
abstract：:Plasmids containing a dihydrofolate reductase (DHFR) expression unit were transfected into DHFR-deficient Chinese hamster ovary (CHO) cells. Methotrexate exposure was used to select cells with amplified DHFR sequences. Three cell lines were isolated containing amplified copies of transfected DNA that had integrated in...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/17.4.1697
更新日期：1989-02-25 00:00:00
abstract：:Histone variants play an important role in shaping the mammalian epigenome and their aberrant expression is frequently observed in several types of cancer. However, the mechanisms that mediate their function and the composition of the variant-containing chromatin are still largely unknown. A proteomic interrogation of...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gku303
更新日期：2014-06-01 00:00:00
abstract：:Ribosome synthesis involves the concomitance of pre-rRNA processing and ribosomal protein assembly. In eukaryotes, this is a complex process that requires the participation of specific sequences and structures within the pre-rRNAs, at least 200 trans-acting factors and the ribosomal proteins. There is little informati...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkq260
更新日期：2010-08-01 00:00:00
abstract：:The majority of genome-wide association study (GWAS) risk variants reside in non-coding DNA sequences. Understanding how these sequence modifications lead to transcriptional alterations and cell-to-cell variability can help unraveling genotype-phenotype relationships. Here, we describe a computational method, dubbed C...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkw1263
更新日期：2017-03-17 00:00:00
abstract：:The Network of Cancer Genes (NCG) collects and integrates data on 736 human genes that are mutated in various types of cancer. For each gene, NCG provides information on duplicability, orthology, evolutionary appearance and topological properties of the encoded protein in a comprehensive version of the human protein-p...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkp957
更新日期：2010-01-01 00:00:00
abstract：:Poly(ADP-ribosyl)ation is involved in numerous bio-logical processes including DNA repair, transcription and cell death. Cellular levels of poly(ADP-ribose) (PAR) are regulated by PAR polymerases (PARPs) and the degrading enzyme PAR glycohydrolase (PARG), controlling the cell fate decision between life and death in re...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gku505
更新日期：2014-07-01 00:00:00
abstract：:A computer program has been developed which aids in the determination of restriction enzyme recognition sequences. This is achieved by cleaving DNAs of known sequence with a restriction endonuclease and comparing the fragmentation pattern with a computer-generated set of patterns. The feasibility of this approach has ...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/5.11.4105
更新日期：1978-11-01 00:00:00
abstract：:Isopentenyl adenosine derivatives are always located adjacent to the 3' end of the anticodon in transfer RNA and have been implicated in certain biological functions. In the enteric bacterium, E. coli, the derivative is ms2i6A whereas in some plant associated bacteria the derivative is the hydroxylated form, ms2io6A. ...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/10.18.5663
更新日期：1982-09-25 00:00:00
abstract：:CLASPIN is an essential mediator in the DNA replication checkpoint, responsible for ATR (ataxia telangiectasia and Rad3-related protein)-dependent activation of CHK1 (checkpoint kinase 1). Here we found a dynamic signaling pathway that regulates CLASPIN turn over. Under unperturbed conditions, the E3 ubiquitin ligase ...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gku978
更新日期：2014-12-01 00:00:00
abstract：:Human lung epithelioid cells were treated with Benzo (a) pyrene diol epoxide (anti) in order to establish the binding and removal of covalent adducts in chromosomal components. Isolating two different classes of mononucleosomes, it was found that their DNA contained different concentrations of B(a)PDE-DNA adducts, whi...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/14.24.9897
更新日期：1986-12-22 00:00:00
abstract：:The emergence of massively parallel sequencing technology has revolutionized microbial profiling, allowing the unprecedented comparison of microbial diversity across time and space in a wide range of host-associated and environmental ecosystems. Although the high-throughput nature of such methods enables the detection...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkt1070
更新日期：2013-12-01 00:00:00
abstract：:We show that in mouse myoblasts the MyoD1 promoter is highly stimulated by MyoD1 expression, suggesting that it is controlled by a positive feedback loop. Using deletion and mutation analyses, we identified the targets for MyoD1 promoter autoregulation as the two proximal E-boxes located close to the MyoD1 core promot...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/22.12.2234
更新日期：1994-06-25 00:00:00
abstract：:Motor proteins that translocate on nucleic acids are key players in gene expression and maintenance. While the function of these proteins is diverse, they are driven by highly conserved core motor domains. In transcription-coupled DNA repair, motor activity serves to remove RNA polymerase stalled on damaged DNA, makin...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkp680
更新日期：2009-10-01 00:00:00
abstract：:Polyadenylation is a co-transcriptional process that modifies mRNA 3'-ends in eukaryotes. In yeast, CF IA and CPF constitute the core 3'-end maturation complex. CF IA comprises Rna14p, Rna15p, Pcf11p and Clp1p. CF IA interacts with the C-terminal domain of RNA Pol II largest subunit via Pcf11p which links pre-mRNA 3'-...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkr800
更新日期：2012-02-01 00:00:00
abstract：:The 279-bp major breakpoint region (mbr) within the 3'-untranslated region (3'-UTR) of the BCL2 gene is a binding site of special AT-rich sequence binding protein 1 (SATB1) that is well known to participate in the long-range regulation of gene transcription. Our previous studies have revealed that the mbr could regula...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkr023
更新日期：2011-06-01 00:00:00
abstract：:The plant gene enod40 is highly conserved among legumes and also present in various non-legume species. It is presumed to play a central regulatory role in the Rhizobium-legume interaction, being expressed well before the initiation of cortical cell divisions resulting in nodule formation. Two small peptides encoded b...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkg721
更新日期：2003-09-01 00:00:00
abstract：:Poly (ADP-ribose) polymerase 1 (PARP1) has emerged as an attractive target for cancer therapy due to its key role in DNA repair processes. Inhibition of PARP1 in BRCA-mutated cancers has been observed to be clinically beneficial. Recent genome-mapping experiments have identified a non-canonical G-quadruplex-forming se...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gky1179
更新日期：2019-02-20 00:00:00
abstract：:A hallmark of defense mechanisms based on clustered regularly interspaced short palindromic repeats (CRISPR) and associated sequences (Cas) are the crRNAs that guide these complexes in the destruction of invading DNA or RNA. Three separate CRISPR-Cas systems exist in the cyanobacterium Synechocystis sp. PCC 6803. Base...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkw786
更新日期：2017-01-25 00:00:00
abstract：:RNA is emerging as a key regulator of a plethora of biological processes. While its study has remained elusive for decades, the recent advent of high-throughput sequencing technologies provided the unique opportunity to develop novel techniques for the study of RNA structure and post-transcriptional modifications. Non...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gky486
更新日期：2018-09-19 00:00:00
abstract：:Genes from yeast to mammals are frequently subject to non-coding transcription of their antisense strand; however the genome-wide role for antisense transcription remains elusive. As transcription influences chromatin structure, we took a genome-wide approach to assess which chromatin features are associated with nasc...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkv666
更新日期：2015-09-18 00:00:00
abstract：:COMPASS is a method for homology detection and local alignment construction based on the comparison of multiple sequence alignments (MSAs). The method derives numerical profiles from given MSAs, constructs local profile-profile alignments and analytically estimates E-values for the detected similarities. Until now, CO...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkm293
更新日期：2007-07-01 00:00:00
abstract：:The PSI/TM-Coffee web server performs multiple sequence alignment (MSA) of proteins by combining homology extension with a consistency based alignment approach. Homology extension is performed with Position Specific Iterative (PSI) BLAST searches against a choice of redundant and non-redundant databases. The main nove...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkw300
更新日期：2016-07-08 00:00:00
abstract：:The complete mitochondrial (mt) genome of Crossostoma lacustre, a freshwater loach from mountain stream of Taiwan, has been cloned and sequenced. This fish mt genome, consisting of 16558 base-pairs, encodes genes for 13 proteins, two rRNAs, and 22 tRNAs, in addition to a regulatory sequence for replication and transcr...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/20.18.4853
更新日期：1992-09-25 00:00:00
abstract：:More than one-third of the RAG1 protein can be truncated from the N-terminus with only subtle effects on the products of V(D)J recombination in vitro or in a mouse. What, then, is the function of the N-terminal domain? We believe it to be regulatory. We determined, several years ago, that an included RING motif could ...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkp192
更新日期：2009-06-01 00:00:00
abstract：:MicroRNAs (miRNAs) are small noncoding RNAs that act as master regulators in many biological processes. miRNAs function mainly by downregulating the expression of their gene targets. Thus, accurate prediction of miRNA targets is critical for characterization of miRNA functions. To this end, we have developed an online...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkz757
更新日期：2020-01-08 00:00:00
abstract：:PIWI-interacting RNAs (piRNAs) comprise a class of small RNAs best known for suppressing transposable elements in germline tissues. The vector mosquito Aedes aegypti encodes seven PIWI genes, four of which are somatically expressed. This somatic piRNA pathway generates piRNAs from viral RNA during infection with cytop...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gky1266
更新日期：2019-03-18 00:00:00
abstract：:When averaged over the full yeast protein-protein interaction and transcriptional regulatory networks, protein hubs with many interaction partners or regulators tend to evolve significantly more slowly due to increased negative selection. However, genome-wide analysis of protein evolution in the subnetworks of associa...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/gkq353
更新日期：2010-10-01 00:00:00
abstract：:Saccharomyces cerevisiae chl1 mutants have a significant increase in the rate of chromosome missegregation. CHL1 encodes a 99 kDa predicted protein with an ATP binding site consensus, a putative helix-turn-helix DNA binding motif, and homology to helicases. Using site-directed mutagenesis, I show that mutations that a...
journal_title：Nucleic acids research
pub_type： 杂志文章
doi：10.1093/nar/28.16.3056
更新日期：2000-08-15 00:00:00
