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https://www.readbyqxmd.com/read/28346890/high-throughput-system-wide-engineering-and-screening-for-microbial-biotechnology
#1
REVIEW
Yannick Vervoort, Alicia Gutiérrez Linares, Miguel Roncoroni, Chengxun Liu, Jan Steensels, Kevin J Verstrepen
Genetic engineering and screening of large number of cells or populations is a crucial bottleneck in today's systems biology and applied (micro)biology. Instead of using standard methods in bottles, flasks or 96-well plates, scientists are increasingly relying on high-throughput strategies that miniaturize their experiments to the nanoliter and picoliter scale and the single-cell level. In this review, we summarize different high-throughput system-wide genome engineering and screening strategies for microbes...
March 24, 2017: Current Opinion in Biotechnology
https://www.readbyqxmd.com/read/28344037/methods-for-decoding-cas9-protospacer-adjacent-motif-pam-sequences-a-brief-overview
#2
REVIEW
Tautvydas Karvelis, Giedrius Gasiunas, Virginijus Siksnys
Recently the Cas9, an RNA guided DNA endonuclease, emerged as a powerful tool for targeted genome manipulations. Cas9 protein can be reprogrammed to cleave, bind or nick any DNA target by simply changing crRNA sequence, however a short nucleotide sequence, termed PAM, is required to initiate crRNA hybridization to the DNA target. PAM sequence is recognized by Cas9 protein and must be determined experimentally for each Cas9 variant. Exploration of Cas9 orthologs could offer a diversity of PAM sequences and novel biochemical properties that may be beneficial for genome editing applications...
March 23, 2017: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28334774/high-tolerance-to-self-targeting-of-the-genome-by-the-endogenous-crispr-cas-system-in-an-archaeon
#3
Aris-Edda Stachler, Israela Turgeman-Grott, Ella Shtifman-Segal, Thorsten Allers, Anita Marchfelder, Uri Gophna
CRISPR-Cas systems allow bacteria and archaea to acquire sequence-specific immunity against selfish genetic elements such as viruses and plasmids, by specific degradation of invader DNA or RNA. However, this involves the risk of autoimmunity if immune memory against host DNA is mistakenly acquired. Such autoimmunity has been shown to be highly toxic in several bacteria and is believed to be one of the major costs of maintaining these defense systems. Here we generated an experimental system in which a non-essential gene, required for pigment production and the reddish colony color, is targeted by the CRISPR-Cas I-B system of the halophilic archaeon Haloferax volcanii...
March 2, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28334141/distribution-of-antimicrobial-resistance-determinants-virulence-associated-factors-and-crispr-loci-in-isolates-of-enterococcus-faecalis-from-various-settings-and-genetic-lineages
#4
Iwona Gawryszewska, Katarzyna Malinowska, Alicja Kuch, Dorota Chrobak-Chmiel, Lucja Laniewska-Trokenheim, Waleria Hryniewicz, Ewa Sadowy
Enterococcus faecalis represents an important factor of hospital-associated infections (HAIs). The knowledge on its evolution from a commensal to an opportunistic pathogen is still limited, thus we performed a study to characterize distribution of factors that may contribute to this adaptation. Using a collection obtained from various settings (hospitalized patients, community carriers, animals, fresh food, sewage, water) we investigated differences in antimicrobial susceptibility, distribution of antimicrobial resistance genes, virulence-associated determinants and phenotypes, and CRISPR loci in the context of the clonal relatedness of isolates...
March 1, 2017: Pathogens and Disease
https://www.readbyqxmd.com/read/28331970/disease-modeling-in-genetic-kidney-diseases-zebrafish
#5
REVIEW
Heiko Schenk, Janina Müller-Deile, Mark Kinast, Mario Schiffer
Growing numbers of translational genomics studies are based on the highly efficient and versatile zebrafish (Danio rerio) vertebrate model. The increasing types of zebrafish models have improved our understanding of inherited kidney diseases, since they not only display pathophysiological changes but also give us the opportunity to develop and test novel treatment options in a high-throughput manner. New paradigms in inherited kidney diseases have been developed on the basis of the distinct genome conservation of approximately 70 % between zebrafish and humans in terms of existing gene orthologs...
March 22, 2017: Cell and Tissue Research
https://www.readbyqxmd.com/read/28329204/new-neuroscience-tools-that-are-identifying-the-sleep-wake-circuit
#6
Priyattam J Shiromani, John H Peever
The complexity of the brain is yielding to technology. In the area of sleep neurobiology, conventional neuroscience tools such as lesions, cell recordings, c-Fos, and axon-tracing methodologies have been instrumental in identifying the complex and intermingled populations of sleep- and arousal-promoting neurons that orchestrate and generate wakefulness, NREM, and REM sleep. In the last decade, new technologies such as optogenetics, chemogenetics and the CRISPR-Cas system have begun to transform how biologists understand the finer details associated with sleep-wake regulation...
March 1, 2017: Sleep
https://www.readbyqxmd.com/read/28327544/fmnl-formins-boost-lamellipodial-force-generation
#7
Frieda Kage, Moritz Winterhoff, Vanessa Dimchev, Jan Mueller, Tobias Thalheim, Anika Freise, Stefan Brühmann, Jana Kollasser, Jennifer Block, Georgi Dimchev, Matthias Geyer, Hans-Joachim Schnittler, Cord Brakebusch, Theresia E B Stradal, Marie-France Carlier, Michael Sixt, Josef Käs, Jan Faix, Klemens Rottner
Migration frequently involves Rac-mediated protrusion of lamellipodia, formed by Arp2/3 complex-dependent branching thought to be crucial for force generation and stability of these networks. The formins FMNL2 and FMNL3 are Cdc42 effectors targeting to the lamellipodium tip and shown here to nucleate and elongate actin filaments with complementary activities in vitro. In migrating B16-F1 melanoma cells, both formins contribute to the velocity of lamellipodium protrusion. Loss of FMNL2/3 function in melanoma cells and fibroblasts reduces lamellipodial width, actin filament density and -bundling, without changing patterns of Arp2/3 complex incorporation...
March 22, 2017: Nature Communications
https://www.readbyqxmd.com/read/28293232/commentary-type-i-crispr-cas-targets-endogenous-genes-and-regulates-virulence-to-evade-mammalian-host-immunity
#8
COMMENT
Hanna Müller-Esparza, Lennart Randau
No abstract text is available yet for this article.
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/28288698/crispr-system-for-genome-engineering-the-application-for-autophagy-study
#9
Jianzhou Cui, Shirley Jia Li Chew, Yin Shi, Zhiyuan Gong, Han-Ming Shen
CRISPR/Cas9 is the latest tool introduced in the field of genome engineering and is so far the best genome-editing tool as compared to its precedents such as, meganucleases, zinc finger nucleases (ZFNs) and transcription activator-like effectors (TALENs). The simple design and assembly of the CRISPR/Cas9 system makes genome editing easy to perform as it uses small guide RNAs that correspond to their DNA targets for high efficiency editing. This has helped open the doors for multiplexible genome targeting in many species that were intractable using old genetic perturbation techniques...
March 14, 2017: BMB Reports
https://www.readbyqxmd.com/read/28287876/crispr-cas9-from-a-bacterial-immune-system-to-genome-edited-human-cells-in-clinical-trials
#10
Leonhard Kick, Marion Kirchner, Sabine Schneider
The adaptive bacterial immune system CRISPR-Cas is revolutionising all fields of life science and has opened up new frontiers towards personalised medicine. Since the elucidation of the molecular mechanism of Cas9 from Streptococcus pyogenes in 2012 and its development as a genomic engineering tool, genetic modifications in more than 40 species have been carried out, over 290 patents have been filed worldwide and the first clinical trials using CRISPR-Cas-modified T-cells have recently been started in China and in the US...
March 13, 2017: Bioengineered
https://www.readbyqxmd.com/read/28284774/analysis-of-microsatellite-instability-in-crispr-cas9-editing-mice
#11
Xueyun Huo, Yating Du, Jing Lu, Meng Guo, Zhenkun Li, Shuangyue Zhang, Xiaohong Li, Zhenwen Chen, Xiaoyan Du
Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR- associated (Cas) protein 9 system is a novel and powerful tool which is widely used for genome editing. CRISPR/Cas9 is RNA-guided and can lead to desired genomic modifications. However, whether the CRISPR/Cas9-mediated genome editing causes genomic alterations and genomic instability, such as microsatellite instability (MSI), is still unknown. Here we detected MSI in 21 CRISPR/Cas9 mouse strains using a panel of 42 microsatellite loci which were selected from our previous studies...
February 28, 2017: Mutation Research
https://www.readbyqxmd.com/read/28272984/exogenous-gene-integration-mediated-by-genome-editing-technologies-in-zebrafish
#12
Hitoshi Morita, Kiyohito Taimatsu, Kanoko Yanagi, Atsuo Kawahara
Genome editing technologies, such as transcription activator-like effector nuclease (TALEN) and the clustered regularly interspaced short palindromic repeat (CRISPR)/ CRISPR-associated protein (Cas) systems, can induce DNA double-strand breaks (DSBs) at the targeted genomic locus, leading to frameshift-mediated gene disruption in the process of DSB repair. Recently, the technology-induced DSBs followed by DSB repairs are applied to integrate exogenous genes into the targeted genomic locus in various model organisms...
March 8, 2017: Bioengineered
https://www.readbyqxmd.com/read/28270274/crispr-cas-system-presents-multiple-transcriptional-units-including-antisense-rnas-that-are-expressed-in-minimal-medium-and-upregulated-by-ph-in-salmonella-enterica-serovar-typhi
#13
Liliana Medina-Aparicio, Javier E Rebollar-Flores, América A Beltrán-Luviano, Alejandra Vázquez, Rosa M Gutiérrez-Ríos, Leticia Olvera, Edmundo Calva, Ismael Hernández-Lucas
The CRISPR-Cas system is involved in bacterial immunity, virulence, gene regulation, biofilm formation and sporulation. In Salmonella enterica serovar Typhi, this system consists of five transcriptional units including antisense RNAs. It was determined that these genetic elements are expressed in minimal medium and are up-regulated by pH. In addition, a transcriptional characterization of cas3 and ascse2-1 is included herein.
February 2017: Microbiology
https://www.readbyqxmd.com/read/28264995/the-dev-operon-regulates-the-timing-of-sporulation-during-myxococcus-xanthus-development
#14
Ramya Rajagopalan, Lee Kroos
Myxococcus xanthus undergoes multicellular development when starved. Thousands of rod-shaped cells coordinate their movements and aggregate into mounds in which cells differentiate into spores. Mutations in the dev operon impair development. The dev operon encompasses a clustered regularly interspaced short palindromic repeat-associated (CRISPR-Cas) system. Null mutations in devI, a small gene at the beginning of the dev operon, suppress the developmental defects caused by null mutations in the downstream devR and devS genes, but failed to suppress defects caused by a small, in-frame deletion in devT We provide evidence that the original mutant has a second-site mutation...
March 6, 2017: Journal of Bacteriology
https://www.readbyqxmd.com/read/28264992/crispr-cas-and-contact-dependent-secretion-systems-present-on-excisable-pathogenicity-islands-with-conserved-recombination-modules
#15
Megan R Carpenter, Sai S Kalburge, Joseph D Borowski, Molly C Peters, Rita R Colwell, E Fidelma Boyd
Pathogenicity islands (PAIs) are mobile integrated genetic elements that contain a diverse range of virulence factors. PAIs integrate into the host chromosome at a tRNA locus that contains their specific bacterial attachment site, attB, via integrase mediated site-specific recombination generating attL and attR sites. We identified conserved recombination modules (integrases and att sites) previously described in choleragenic V. cholerae PAIs but with novel cargo genes. Clustered regularly interspaced short palindromic repeat (CRISPR)-associated proteins (Cas proteins) and a type VI secretion system (T6SS) gene cluster were identified at the Vibrio Pathogenicity Island-1 (VPI-1) insertion site in nineteen V...
March 6, 2017: Journal of Bacteriology
https://www.readbyqxmd.com/read/28264649/segmental-duplications-and-evolutionary-acquisition-of-uv-damage-response-in-the-spata31-gene-family-of-primates-and-humans
#16
Cemalettin Bekpen, Sven Künzel, Chen Xie, Muthukrishnan Eaaswarkhanth, Yen-Lung Lin, Omer Gokcumen, Cezmi A Akdis, Diethard Tautz
BACKGROUND: Segmental duplications are an abundant source for novel gene functions and evolutionary adaptations. This mechanism of generating novelty was very active during the evolution of primates particularly in the human lineage. Here, we characterize the evolution and function of the SPATA31 gene family (former designation FAM75A), which was previously shown to be among the gene families with the strongest signal of positive selection in hominoids. The mouse homologue for this gene family is a single copy gene expressed during spermatogenesis...
March 6, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28258551/crispr-cas9-mediated-targeted-mutagenesis-in-upland-cotton-gossypium-hirsutum-l
#17
Madhusudhana R Janga, LeAnne M Campbell, Keerti S Rathore
The clustered, regularly interspaced, short palindromic repeats (CRISPR)/CRISPR associated (Cas)9 protein system has emerged as a simple and efficient tool for genome editing in eukaryotic cells. It has been shown to be functional in several crop species, yet there are no reports on the application of this or any other genome editing technologies in the cotton plant. Cotton is an important crop that is grown mainly for its fiber, but its seed also serves as a useful source of edible oil and feed protein. Most of the commercially-grown cotton is tetraploid, thus making it much more difficult to target both sets of homeologous alleles...
March 3, 2017: Plant Molecular Biology
https://www.readbyqxmd.com/read/28254804/gene-editing-with-crispr-cas9-rna-directed-nuclease
#18
REVIEW
Thomas Doetschman, Teodora Georgieva
Genetic engineering of model organisms and cultured cells has for decades provided important insights into the mechanisms underlying cardiovascular development and disease. In the past few years the development of several nuclease systems has broadened the range of model/cell systems that can be engineered. Of these, the CRISPR (clustered regularly interspersed short palindromic repeats)/Cas9 (CRISPR-associated protein 9) system has become the favorite for its ease of application. Here we will review this RNA-guided nuclease system for gene editing with respect to its usefulness for cardiovascular studies and with an eye toward potential therapy...
March 3, 2017: Circulation Research
https://www.readbyqxmd.com/read/28247934/anti-crispr-proteins-counterattack-of-phages-on-bacterial-defense-crispr-cas-system
#19
Kulbhushan Chaudhary, Anirudha Chattopadhyay, Dharmendra Pratap
Since the dawn of life there is a never ending strife between bacteria and phages. Both are perpetually changing their strategies to take over each other. CRISPR/Cas is the most widespread defense system used by bacteria against mobile genetic elements (MGEs) such as phages, cojugative palsmids, transoposons and pathogenicity islands. This system utilizes small guide RNA molecules to protect against phages infection and invasion by MGEs. Phages circumvent to these antiviral barriers by point mutation in PAM (protospacer-adjacent motif) sequence, genome rearrangements and by using anti-CRISPR proteins...
March 1, 2017: Journal of Cellular Physiology
https://www.readbyqxmd.com/read/28246397/relationship-between-somatic-mosaicism-of-pax6-mutation-and-variable-developmental-eye-abnormalities-an-analysis-of-crispr-genome-edited-mouse-embryos
#20
Akihiro Yasue, Hitomi Kono, Munenori Habuta, Tetsuya Bando, Keita Sato, Junji Inoue, Seiichi Oyadomari, Sumihare Noji, Eiji Tanaka, Hideyo Ohuchi
The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein (Cas) system is a rapid gene-targeting technology that does not require embryonic stem cells. To demonstrate dosage effects of the Pax6 gene on eye formation, we generated Pax6-deficient mice with the CRISPR/Cas system. Eyes of founder embryos at embryonic day (E) 16.5 were examined and categorized according to macroscopic phenotype as class 1 (small eye with distinct pigmentation), class 2 (pigmentation without eye globes), or class 3 (no pigmentation and no eyes)...
December 2017: Scientific Reports
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