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Crispr-Cas

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https://www.readbyqxmd.com/read/29228332/prespacer-processing-and-specific-integration-in-a-type-i-a-crispr-system
#1
Clare Rollie, Shirley Graham, Christophe Rouillon, Malcolm F White
The CRISPR-Cas system for prokaryotic adaptive immunity provides RNA-mediated protection from viruses and mobile genetic elements. Adaptation is dependent on the Cas1 and Cas2 proteins along with varying accessory proteins. Here we analyse the process in Sulfolobus solfataricus, showing that while Cas1 and Cas2 catalyze spacer integration in vitro, host factors are required for specificity. Specific integration also requires at least 400 bp of the leader sequence, and is dependent on the presence of hydrolysable ATP, suggestive of an active process that may involve DNA remodelling...
December 8, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29222508/crispr-cpf1-mediates-efficient-homology-directed-repair-and-temperature-controlled-genome-editing
#2
Miguel A Moreno-Mateos, Juan P Fernandez, Romain Rouet, Charles E Vejnar, Maura A Lane, Emily Mis, Mustafa K Khokha, Jennifer A Doudna, Antonio J Giraldez
Cpf1 is a novel class of CRISPR-Cas DNA endonucleases, with a wide range of activity across different eukaryotic systems. Yet, the underlying determinants of this variability are poorly understood. Here, we demonstrate that LbCpf1, but not AsCpf1, ribonucleoprotein complexes allow efficient mutagenesis in zebrafish and Xenopus. We show that temperature modulates Cpf1 activity by controlling its ability to access genomic DNA. This effect is stronger on AsCpf1, explaining its lower efficiency in ectothermic organisms...
December 8, 2017: Nature Communications
https://www.readbyqxmd.com/read/29218031/crispr-cas-systems-in-bacteroides-fragilis-an-important-pathobiont-in-the-human-gut-microbiome
#3
Mehrdad Tajkarimi, Hannah M Wexler
Background: While CRISPR-Cas systems have been identified in bacteria from a wide variety of ecological niches, there are no studies to describe CRISPR-Cas elements in Bacteroides species, the most prevalent anaerobic bacteria in the lower intestinal tract. Microbes of the genus Bacteroides make up ~25% of the total gut microbiome. Bacteroides fragilis comprises only 2% of the total Bacteroides in the gut, yet causes of >70% of Bacteroides infections. The factors causing it to transition from benign resident of the gut microbiome to virulent pathogen are not well understood, but a combination of horizontal gene transfer (HGT) of virulence genes and differential transcription of endogenous genes are clearly involved...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/29216382/improving-crispr-cas-specificity-with-chemical-modifications-in-single-guide-rnas
#4
Daniel E Ryan, David Taussig, Israel Steinfeld, Smruti M Phadnis, Benjamin D Lunstad, Madhurima Singh, Xuan Vuong, Kenji D Okochi, Ryan McCaffrey, Magdalena Olesiak, Subhadeep Roy, Chong Wing Yung, Bo Curry, Jeffrey R Sampson, Laurakay Bruhn, Douglas J Dellinger
CRISPR systems have emerged as transformative tools for altering genomes in living cells with unprecedented ease, inspiring keen interest in increasing their specificity for perfectly matched targets. We have developed a novel approach for improving specificity by incorporating chemical modifications in guide RNAs (gRNAs) at specific sites in their DNA recognition sequence ('guide sequence') and systematically evaluating their on-target and off-target activities in biochemical DNA cleavage assays and cell-based assays...
December 4, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29216246/draft-genome-sequence-of-actinotignum-schaalii-dsm-15541t-genetic-insights-into-the-lifestyle-cell-fitness-and-virulence
#5
Atteyet F Yassin, Stefan Langenberg, Marcel Huntemann, Alicia Clum, Manoj Pillay, Krishnaveni Palaniappan, Neha Varghese, Natalia Mikhailova, Supratim Mukherjee, T B K Reddy, Chris Daum, Nicole Shapiro, Natalia Ivanova, Tanja Woyke, Nikos C Kyrpides
The permanent draft genome sequence of Actinotignum schaalii DSM 15541T is presented. The annotated genome includes 2,130,987 bp, with 1777 protein-coding and 58 rRNA-coding genes. Genome sequence analysis revealed absence of genes encoding for: components of the PTS systems, enzymes of the TCA cycle, glyoxylate shunt and gluconeogensis. Genomic data revealed that A. schaalii is able to oxidize carbohydrates via glycolysis, the nonoxidative pentose phosphate and the Entner-Doudoroff pathways. Besides, the genome harbors genes encoding for enzymes involved in the conversion of pyruvate to lactate, acetate and ethanol, which are found to be the end products of carbohydrate fermentation...
2017: PloS One
https://www.readbyqxmd.com/read/29212058/genetic-analysis-and-crispr-typing-of-salmonella-enterica-serovar-enteritidis-from-different-sources-revealed-potential-transmission-from-poultry-and-pig-to-human
#6
Qiuchun Li, Xin Wang, Kequan Yin, Yachen Hu, Haiyan Xu, Xiaolei Xie, Lijuan Xu, Xiao Fei, Xiang Chen, Xinan Jiao
Salmonella enterica serovar Enteritidis (S. Enteritidis) is one of the most prevalent serotypes in Salmonella isolated from poultry and the most commonly reported cause of human salmonellosis. In this study, we aimed to assess the genetic diversity of 329 S. Enteritidis strains isolated from different sources from 2009 to 2016 in China. Clustered regularly interspaced short palindromic repeat (CRISPR) typing was used to characterize these 262 chicken clinical isolates, 38 human isolates, 18 pig isolates, six duck isolates, three goose isolates and two isolates of unknown source...
November 28, 2017: International Journal of Food Microbiology
https://www.readbyqxmd.com/read/29205229/crispr-cas-mediated-gene-silencing-reveals-racr-to-be-a-negative-regulator-of-ydas-and-ydat-toxins-in-escherichia-coli-k-12
#7
Gargi Bindal, Revathy Krishnamurthi, Aswin Sai Narain Seshasayee, Devashish Rath
Bacterial genomes are rich in horizontally acquired prophages. racR is an essential gene located in the rac prophage that is resident in many Escherichia coli genomes. Employing a clustered regularly interspaced short palindromic repeat (CRISPR)-Cas-based gene silencing approach, we show that RacR is a negative regulator of the divergently transcribed and adjacent ydaS-ydaT operon in Escherichia coli K-12. Overexpression of YdaS and YdaT due to RacR depletion leads to cell division defects and decrease in survival...
November 2017: MSphere
https://www.readbyqxmd.com/read/29202216/type-ii-c-crispr-cas9-biology-mechanism-and-application
#8
Aamir Mir, Alireza Edraki, Jooyoung Lee, Erik J Sontheimer
Genome editing technologies have been revolutionized by the discovery of prokaryotic RNA-guided defense system called CRISPR-Cas. Cas9, a single effector protein found in type II CRISPR systems, has been at the heart of this genome editing revolution. Nearly half of the Cas9s discovered so far belong to the type II-C subtype, but have not been explored extensively. Type II-C CRISPR-Cas systems are the simplest of the type II systems, employing only three Cas proteins. Cas9s are central players in type II-C systems since they function in multiple steps of the CRISPR pathway, including adaptation and interference...
December 4, 2017: ACS Chemical Biology
https://www.readbyqxmd.com/read/29202180/emergence-and-spread-of-epidemic-multidrug-resistant-pseudomonas-aeruginosa
#9
Tohru Miyoshi-Akiyama, Tatsuya Tada, Norio Ohmagari, Nguyen Viet Hung, Prasit Tharavichitkul, Bharat Mani Pokhrel, Marek Gniadkowski, Masahiro Shimojima, Teruo Kirikae
Pseudomonas aeruginosa (P. aeruginosa) is one of the most common nosocomial pathogens worldwide. Although the emergence of multidrug-resistant (MDR) P. aeruginosa is a critical problem in medical practice, the key features involved in the emergence and spread of MDR P. aeruginosa remain unknown. This study utilized whole genome sequence (WGS) analyses to define the population structure of 185 P. aeruginosa clinical isolates from several countries. Of these 185 isolates, 136 were categorized into sequence type (ST) 235, one of the most common types worldwide...
November 29, 2017: Genome Biology and Evolution
https://www.readbyqxmd.com/read/29197619/harnessing-crispr-cas-systems-for-programmable-transcriptional-and-post-transcriptional-regulation
#10
REVIEW
Ahmed Mahas, C Neal Stewart, Magdy M Mahfouz
Genome editing has enabled broad advances and novel approaches in studies of gene function and structure; now, emerging methods aim to precisely engineer post-transcriptional processes. Developing precise, efficient molecular tools to alter the transcriptome holds great promise for biotechnology and synthetic biology applications. Different approaches have been employed for targeted degradation of RNA species in eukaryotes, but they lack programmability and versatility, thereby limiting their utility for diverse applications...
November 29, 2017: Biotechnology Advances
https://www.readbyqxmd.com/read/29189942/the-conspicuity-of-crispr-cpf1-system-as-a-significant-breakthrough-in-genome-editing
#11
REVIEW
Hadi Bayat, Mohammad Hossein Modarressi, Azam Rahimpour
Clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein (Cas) is a microbial adaptive immune system. CRISPR-Cas systems are classified into two main classes and six types. Cpf1 is a putative type V (class II) CRISPR effector, which has revolutionized the genome editing approaches through multiple distinct features such as using T-rich protospacer-adjacent motif, applying a short guide RNA lacking trans-activating crRNA, introducing a staggered double-strand break, and possessing RNA processing activity in addition to DNA nuclease activity...
November 30, 2017: Current Microbiology
https://www.readbyqxmd.com/read/29188510/methods-for-in-vivo-crispr-cas-editing-of-the-adult-murine-retina
#12
Sandy S Hung, Fan Li, Jiang-Hui Wang, Anna E King, Bang V Bui, Guei-Sheung Liu, Alex W Hewitt
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated protein (Cas) is used by some bacteria and most archaea to protect against viral phage intrusion and has recently been adapted to allow for efficient editing of the mammalian genome. Whilst CRISPR/Cas-based technology has been used to modify genes in mammalian cells in vitro, delivery of CRISPR/Cas system into mammalian tissue and/or organs is more difficult and often requires additional vectors. With the use of adeno-associated virus (AAV) gene delivery system, active CRISPR/Cas enzyme can be maintained for an extended period of time and enable efficient editing of genome in the retina in vivo...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29182776/crystal-structure-of-an-anti-crispr-protein-acriia1
#13
Donghyun Ka, So Young An, Jeong-Yong Suh, Euiyoung Bae
Clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins provide bacteria with RNA-based adaptive immunity against phage infection. To counteract this defense mechanism, phages evolved anti-CRISPR (Acr) proteins that inactivate the CRISPR-Cas systems. AcrIIA1, encoded by Listeria monocytogenes prophages, is the most prevalent among the Acr proteins targeting type II-A CRISPR-Cas systems and has been used as a marker to identify other Acr proteins. Here, we report the crystal structure of AcrIIA1 and its RNA-binding affinity...
November 22, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29180373/generation-of-bacteriophage-insensitive-mutants-of-streptococcus-thermophilus-using-an-antisense-rna-crispr-cas-silencing-approach
#14
Brian McDonnell, Jennifer Mahony, Laurens Hanemaaijer, Thijs R H M Kouwen, Douwe van Sinderen
Predation of starter lactic acid bacteria such as Streptococcus thermophilus by bacteriophages is a persistent and costly problem in the dairy industry. CRISPR-mediated Bacteriophage Insensitive Mutants (BIMs), while straightforward to generate and verify, can quickly be overcome by mutant phages. The aim of this study was to develop a tool allowing the generation of derivatives of commercial S. thermophilus strains which are resistant to phage attack through a non-CRISPR-mediated mechanism, with the objective of generating BIMs exhibiting stable resistance against a range of isolated lytic S...
November 27, 2017: Applied and Environmental Microbiology
https://www.readbyqxmd.com/read/29179671/phylogenomics-of-cas4-family-nucleases
#15
Sanjarbek Hudaiberdiev, Sergey Shmakov, Yuri I Wolf, Michael P Terns, Kira S Makarova, Eugene V Koonin
BACKGROUND: The Cas4 family endonuclease is a component of the adaptation module in many variants of CRISPR-Cas adaptive immunity systems. Unlike most of the other Cas proteins, Cas4 is often encoded outside CRISPR-cas loci (solo-Cas4) and is also found in mobile genetic elements (MGE-Cas4). RESULTS: As part of our ongoing investigation of CRISPR-Cas evolution, we explored the phylogenomics of the Cas4 family. About 90% of the archaeal genomes encode Cas4 compared to only about 20% of the bacterial genomes...
November 28, 2017: BMC Evolutionary Biology
https://www.readbyqxmd.com/read/29177012/crispr-like-sequences-in-helicobacter-pylori-and-application-in-genotyping
#16
Khotchawan Bangpanwimon, Jaksin Sottisuporn, Pimonsri Mittraparp-Arthorn, Warattaya Ueaphatthanaphanich, Attapon Rattanasupar, Christine Pourcel, Varaporn Vuddhakul
Background: Many bacteria and archaea possess a defense system called clustered regularly interspaced short palindromic repeats (CRISPR) associated proteins (CRISPR-Cas system) against invaders such as phages or plasmids. This system has not been demonstrated in Helicobacter pylori. The numbers of spacer in CRISPR array differ among bacterial strains and can be used as a genetic marker for bacterial typing. Results: A total of 36 H. pylori isolates were collected from patients in three hospitals located in the central (PBH) and southern (SKH) regions of Thailand...
2017: Gut Pathogens
https://www.readbyqxmd.com/read/29174768/acetylation-of-tbx5-by-kat2b-and-kat2a-regulates-heart-and-limb-development
#17
Tushar K Ghosh, José J Aparicio-Sánchez, Sarah Buxton, Ami Ketley, Tasabeeh Mohamed, Catrin S Rutland, Siobhan Loughna, J David Brook
TBX5 plays a critical role in heart and forelimb development. Mutations in TBX5 cause Holt-Oram syndrome, an autosomal dominant condition that affects the formation of the heart and upper-limb. Several studies have provided significant insight into the role of TBX5 in cardiogenesis; however, how TBX5 activity is regulated by other factors is still unknown. Here we report that histone acetyltransferases KAT2A and KAT2B associate with TBX5 and acetylate it at Lys339. Acetylation potentiates its transcriptional activity and is required for nuclear retention...
November 22, 2017: Journal of Molecular and Cellular Cardiology
https://www.readbyqxmd.com/read/29173172/therapeutic-applications-of-crispr-cas-for-duchenne-muscular-dystrophy
#18
Tatianna Wai Ying Wong, Ronald D Cohn
BACKGROUND: Duchenne muscular dystrophy (DMD) is an X-linked neuromuscular disease caused by the lack of dystrophin due to mutations in the DMD gene. Since dystrophin is essential in maintaining the integrity of the sarcolemmal membrane, the absence of the protein leads to muscle damage and DMD disease manifestation. Currently there is no cure with only symptomatic management available. OBJECTIVE: The most recent advancements in DMD therapies do not provide a permanent treatment for DMD...
November 21, 2017: Current Gene Therapy
https://www.readbyqxmd.com/read/29173171/new-developments-in-crispr-cas-based-functional-genomics-and-their-implications-for-research-using-zebrafish
#19
Sergey V Prykhozhij, Lucia Caceres, Jason N Berman
Genome editing using CRISPR/Cas9 has advanced very rapidly in its scope, versatility and ease of use. Zebrafish (Danio rerio) has been one of the vertebrate model species where CRISPR/Cas9 has been applied very extensively for many different purposes and with great success. In particular, disease modeling in zebrafish is useful for testing specific gene variants for pathogenicity in a preclinical setting. Here we describe multiple advances in diverse species and systems that can improve genome editing in zebrafish...
November 21, 2017: Current Gene Therapy
https://www.readbyqxmd.com/read/29170279/multiplex-recording-of-cellular-events-over-time-on-crispr-biological-tape
#20
Ravi U Sheth, Sung Sun Yim, Felix L Wu, Harris H Wang
While dynamics underlie many biological processes, our ability to robustly and accurately profile time-varying biological signals and regulatory programs remains limited. Here, we describe a framework to store temporal biological information directly into the genomes of a cell population. A "biological tape recorder" is developed in which biological signals trigger intracellular DNA production that is then recorded by the CRISPR-Cas adaptation system. This approach enables stable recording over multiple days and accurate reconstruction of temporal and lineage information by sequencing CRISPR arrays...
November 23, 2017: Science
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