Bibliografias temáticas / CRISPRko Screening

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Literatura científica selecionada sobre o tema "CRISPRko Screening"

Autor: Grafiati

Publicado: 1 de abril de 2023

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Artigos de revistas sobre o assunto "CRISPRko Screening"

Evers, Bastiaan, Katarzyna Jastrzebski, Jeroen P. M. Heijmans, Wipawadee Grernrum, Roderick L. Beijersbergen e Rene Bernards. "CRISPR knockout screening outperforms shRNA and CRISPRi in identifying essential genes". Nature Biotechnology 34, n.º 6 (25 de abril de 2016): 631–33. http://dx.doi.org/10.1038/nbt.3536.

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Watters, Kyle E., Christof Fellmann, Hua B. Bai, Shawn M. Ren e Jennifer A. Doudna. "Systematic discovery of natural CRISPR-Cas12a inhibitors". Science 362, n.º 6411 (6 de setembro de 2018): 236–39. http://dx.doi.org/10.1126/science.aau5138.

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Cas12a (Cpf1) is a CRISPR-associated nuclease with broad utility for synthetic genome engineering, agricultural genomics, and biomedical applications. Although bacteria harboring CRISPR-Cas9 or CRISPR-Cas3 adaptive immune systems sometimes acquire mobile genetic elements encoding anti-CRISPR proteins that inhibit Cas9, Cas3, or the DNA-binding Cascade complex, no such inhibitors have been found for CRISPR-Cas12a. Here we use a comprehensive bioinformatic and experimental screening approach to identify three different inhibitors that block or diminish CRISPR-Cas12a–mediated genome editing in human cells. We also find a widespread connection between CRISPR self-targeting and inhibitor prevalence in prokaryotic genomes, suggesting a straightforward path to the discovery of many more anti-CRISPRs from the microbial world.

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Selle, Kurt, Todd R. Klaenhammer e Rodolphe Barrangou. "CRISPR-based screening of genomic island excision events in bacteria". Proceedings of the National Academy of Sciences 112, n.º 26 (15 de junho de 2015): 8076–81. http://dx.doi.org/10.1073/pnas.1508525112.

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Genomic analysis ofStreptococcus thermophilusrevealed that mobile genetic elements (MGEs) likely contributed to gene acquisition and loss during evolutionary adaptation to milk. Clustered regularly interspaced short palindromic repeats–CRISPR-associated genes (CRISPR-Cas), the adaptive immune system in bacteria, limits genetic diversity by targeting MGEs including bacteriophages, transposons, and plasmids. CRISPR-Cas systems are widespread in streptococci, suggesting that the interplay between CRISPR-Cas systems and MGEs is one of the driving forces governing genome homeostasis in this genus. To investigate the genetic outcomes resulting from CRISPR-Cas targeting of integrated MGEs,in silicoprediction revealed four genomic islands without essential genes in lengths from 8 to 102 kbp, totaling 7% of the genome. In this study, the endogenous CRISPR3 type II system was programmed to target the four islands independently through plasmid-based expression of engineered CRISPR arrays. TargetinglacZwithin the largest 102-kbp genomic island was lethal to wild-type cells and resulted in a reduction of up to 2.5-log in the surviving population. Genotyping of Lac−survivors revealed variable deletion events between the flanking insertion-sequence elements, all resulting in elimination of the Lac-encoding island. Chimeric insertion sequence footprints were observed at the deletion junctions after targeting all of the four genomic islands, suggesting a common mechanism of deletion via recombination between flanking insertion sequences. These results established that self-targeting CRISPR-Cas systems may direct significant evolution of bacterial genomes on a population level, influencing genome homeostasis and remodeling.

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Kampmann, Martin, Max A. Horlbeck, Yuwen Chen, Jordan C. Tsai, Michael C. Bassik, Luke A. Gilbert, Jacqueline E. Villalta et al. "Next-generation libraries for robust RNA interference-based genome-wide screens". Proceedings of the National Academy of Sciences 112, n.º 26 (15 de junho de 2015): E3384—E3391. http://dx.doi.org/10.1073/pnas.1508821112.

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Genetic screening based on loss-of-function phenotypes is a powerful discovery tool in biology. Although the recent development of clustered regularly interspaced short palindromic repeats (CRISPR)-based screening approaches in mammalian cell culture has enormous potential, RNA interference (RNAi)-based screening remains the method of choice in several biological contexts. We previously demonstrated that ultracomplex pooled short-hairpin RNA (shRNA) libraries can largely overcome the problem of RNAi off-target effects in genome-wide screens. Here, we systematically optimize several aspects of our shRNA library, including the promoter and microRNA context for shRNA expression, selection of guide strands, and features relevant for postscreen sample preparation for deep sequencing. We present next-generation high-complexity libraries targeting human and mouse protein-coding genes, which we grouped into 12 sublibraries based on biological function. A pilot screen suggests that our next-generation RNAi library performs comparably to current CRISPR interference (CRISPRi)-based approaches and can yield complementary results with high sensitivity and high specificity.

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Göttl, Vanessa L., Ina Schmitt, Kristina Braun, Petra Peters-Wendisch, Volker F. Wendisch e Nadja A. Henke. "CRISPRi-Library-Guided Target Identification for Engineering Carotenoid Production by Corynebacterium glutamicum". Microorganisms 9, n.º 4 (24 de março de 2021): 670. http://dx.doi.org/10.3390/microorganisms9040670.

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Corynebacterium glutamicum is a prominent production host for various value-added compounds in white biotechnology. Gene repression by dCas9/clustered regularly interspaced short palindromic repeats (CRISPR) interference (CRISPRi) allows for the identification of target genes for metabolic engineering. In this study, a CRISPRi-based library for the repression of 74 genes of C. glutamicum was constructed. The chosen genes included genes encoding enzymes of glycolysis, the pentose phosphate pathway, and the tricarboxylic acid cycle, regulatory genes, as well as genes of the methylerythritol phosphate and carotenoid biosynthesis pathways. As expected, CRISPRi-mediated repression of the carotenogenesis repressor gene crtR resulted in increased pigmentation and cellular content of the native carotenoid pigment decaprenoxanthin. CRISPRi screening identified 14 genes that affected decaprenoxanthin biosynthesis when repressed. Carotenoid biosynthesis was significantly decreased upon CRISPRi-mediated repression of 11 of these genes, while repression of 3 genes was beneficial for decaprenoxanthin production. Largely, but not in all cases, deletion of selected genes identified in the CRISPRi screen confirmed the pigmentation phenotypes obtained by CRISPRi. Notably, deletion of pgi as well as of gapA improved decaprenoxanthin levels 43-fold and 9-fold, respectively. The scope of the designed library to identify metabolic engineering targets, transfer of gene repression to stable gene deletion, and limitations of the approach were discussed.

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GÜLER KARA, Hale, Buket KOSOVA, Eda DOĞAN, Vildan BOZOK ÇETİNTAŞ e Şerif ŞENTÜRK. "CRISPR-Cas Functional Genetic Screening: Traditional Review". Turkiye Klinikleri Journal of Medical Sciences 42, n.º 4 (2022): 311–22. http://dx.doi.org/10.5336/medsci.2022-88507.

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Lanning, Bryan R., e Christopher R. Vakoc. "Single-minded CRISPR screening". Nature Biotechnology 35, n.º 4 (abril de 2017): 339–40. http://dx.doi.org/10.1038/nbt.3849.

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Haswell, Jeffrey R., Kaia Mattioli, Chiara Gerhardinger, Philipp G. Maass, Daniel J. Foster, Paola Peinado, Xiaofeng Wang, Pedro P. Medina, John L. Rinn e Frank J. Slack. "Genome-wide CRISPR interference screen identifies long non-coding RNA loci required for differentiation and pluripotency". PLOS ONE 16, n.º 11 (3 de novembro de 2021): e0252848. http://dx.doi.org/10.1371/journal.pone.0252848.

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Although many long non-coding RNAs (lncRNAs) exhibit lineage-specific expression, the vast majority remain functionally uncharacterized in the context of development. Here, we report the first described human embryonic stem cell (hESC) lines to repress (CRISPRi) or activate (CRISPRa) transcription during differentiation into all three germ layers, facilitating the modulation of lncRNA expression during early development. We performed an unbiased, genome-wide CRISPRi screen targeting thousands of lncRNA loci expressed during endoderm differentiation. While dozens of lncRNA loci were required for proper differentiation, most differentially expressed lncRNAs were not, supporting the necessity for functional screening instead of relying solely on gene expression analyses. In parallel, we developed a clustering approach to infer mechanisms of action of lncRNA hits based on a variety of genomic features. We subsequently identified and validated FOXD3-AS1 as a functional lncRNA essential for pluripotency and differentiation. Taken together, the cell lines and methodology described herein can be adapted to discover and characterize novel regulators of differentiation into any lineage.

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Ancos-Pintado, Raquel, Irene Bragado-García, María Luz Morales, Roberto García-Vicente, Andrés Arroyo-Barea, Alba Rodríguez-García, Joaquín Martínez-López, María Linares e María Hernández-Sánchez. "High-Throughput CRISPR Screening in Hematological Neoplasms". Cancers 14, n.º 15 (25 de julho de 2022): 3612. http://dx.doi.org/10.3390/cancers14153612.

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CRISPR is becoming an indispensable tool in biological research, revolutionizing diverse fields of medical research and biotechnology. In the last few years, several CRISPR-based genome-targeting tools have been translated for the study of hematological neoplasms. However, there is a lack of reviews focused on the wide uses of this technology in hematology. Therefore, in this review, we summarize the main CRISPR-based approaches of high throughput screenings applied to this field. Here we explain several libraries and algorithms for analysis of CRISPR screens used in hematology, accompanied by the most relevant databases. Moreover, we focus on (1) the identification of novel modulator genes of drug resistance and efficacy, which could anticipate relapses in patients and (2) new therapeutic targets and synthetic lethal interactions. We also discuss the approaches to uncover novel biomarkers of malignant transformations and immune evasion mechanisms. We explain the current literature in the most common lymphoid and myeloid neoplasms using this tool. Then, we conclude with future directions, highlighting the importance of further gene candidate validation and the integration and harmonization of the data from CRISPR screening approaches.

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Serebrenik, Yevgeniy V., e Ophir Shalem. "CRISPR mutagenesis screening of mice". Nature Cell Biology 20, n.º 11 (8 de outubro de 2018): 1235–37. http://dx.doi.org/10.1038/s41556-018-0224-y.

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Teses / dissertações sobre o assunto "CRISPRko Screening"

Erard, Nicolas Pascal Jean. "Optimization of molecular tools for high-throughput genetic screening". Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/271895.

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Forward genetic screening allows for the identification of any genes important for a particular biological process or phenotype. While the power of this approach is broadly agreed on, the efficacy of currently available tools limits the strength of conclusions drawn from these experiments. This thesis describes a method to optimize molecular tools for high-throughput screening, both for shRNA and sgRNA based reagents. Using large shRNA efficacy datasets, we first designed an algorithm predicting the potency of shRNAs based on sequence determinants. Combined with a novel shRNA backbone that further improves the processing of synthetic shRNAs, we built a library of potent shRNAs to reliably and efficiently knock-down any gene in the human and mouse genomes. We then went on to apply a similar approach to identify sgRNAs with increased activity. We complemented this with conservation and repair prediction to increase the likelihood of generating functional knock-outs. With these tools in hand, we constructed, sequence-verified and validated arrayed shRNA and sgRNA libraries targeting any protein coding gene in the human genome. These resources allow large-scale screens to be performed in a multiplexed or arrayed format in a variety of biological contexts. I have also applied these tools to identify therapeutic targets to circumvent cancer resistance to treatment in two different contexts. To overcome the shortfalls of single target therapy, I have developed multiplexed multidimensional shRNA screening strategy, where two genes are knocked down simultaneously in each cell. This strategy allows the identification of gene pairs that could be targeted in tandem to maximize therapeutic benefits. As a proof of concept, I have used it with a subset of druggable genes in melanoma cell lines. Moreover, we have applied our genome wide shRNA libraries to a different resistance context, stroma-mediated resistance to gemcitabine in PDAC. In this project, we performed screens in a PDAC-CAF coculture setting to try and identify cancer vulnerabilities specifically in the presence of stroma. Overall, the tools developed in this thesis allow for the efficient knockdown or knockout of any gene, both in an individual or combinatorial setting. Apart from providing a resource that will be useful for many fields, we have performed several proof-of-concept studies where we have applied our tools to identify potential cancer drug targets.

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Sheel, Ankur. "Identification of Essential Genes in Hepatocellular Carcinomas using CRISPR Screening". eScholarship@UMMS, 2019. https://escholarship.umassmed.edu/gsbs_diss/1039.

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Hepatocellular carcinoma (HCC) is an aggressive subtype of liver cancer with a poor prognosis. Currently, prognosis for HCC patients remains poor as few therapies are available. The clinical need for more effective HCC treatments remains unmet partially because HCC is genetically heterogeneous and HCC driver genes amenable to targeted therapy are largely unknown. Mutations in the TP53 gene are found in ~30% of HCC patients and confer poor prognosis to patients. Identifying genes whose depletion can inhibit HCC growth, and determining the mechanisms involved, will aid the development of targeted therapies for HCC patients. Therefore, the first half of this thesis focuses on identifying genes that are required for cell growth in HCC independent of p53 status. We performed a kinome-wide CRISPR screen to identify genes required for cell growth in three HCC cell lines: HepG2 (p53 wild-type), Huh7 (p53-mutant) and Hep3B (p53-null) cells. The kinome screen identified 31 genes that were required for cell growth in 3 HCC cell lines independent of TP53 status. Among the 31 genes, 8 genes were highly expressed in HCC compared to normal tissue and increased expression was associated with poor survival in HCC patients. We focused on TRRAP, a co-factor for histone acetyltransferases. TRRAP function has not been previously characterized in HCC. CRISPR/Cas9 mediated depletion of TRRAP reduced cell growth and colony formation in all three cell lines. Moreover, depletion of TRRAP reduced its histone acetyltransferase co-factors KAT2A and KAT5 at the protein level with no change at the mRNA level. I found that depletion of KAT5, but not KAT2A, reduced cell growth. Notably, inhibition of proteasome- and lysosome-mediated degradation failed to rescue protein levels of KAT2A and KAT5 in the absence of TRRAP. Moreover, tumor initiation in an HCC mouse model failed after CRISPR/Cas9 depletion of TRRAP due to clearance via macrophages and HCC cells depleted of TRRAP and KAT5 failed to grow as subcutaneous xenografts in vivo. RNA-seq and bioinformatic analysis of HCC patient samples revealed that TRRAP positively regulates expression of genes that are involved in mitotic progression. In HCC, this subset of genes is clinically relevant as they are overexpressed compared to normal tissue and high expression confers poor survival to patients. I identified TOP2A as one of the mitotic gene targets of the TRRAP/KAT5 complex whose inhibition greatly reduces proliferation of HCC cells. Given that this was the first time the TRRAP/KAT5 complex has been identified as a therapeutic target in HCC, the second half of this thesis focuses on identifying the mechanism via which depletion of this complex inhibits proliferation of HCC cells. I discovered that depletion of TRRAP, KAT5 and TOP2A reduced proliferation of HCC cells by inducing senescence. Typically, senescence is an irreversible state of cell cycle arrest at G1 that is due to activation of p53/p21 expression, phosphorylation of RB, and DNA damage. Surprisingly, induction of senescence after loss of TRRAP, KAT5 and TOP2A arrested cells during G2/M and senescence was independent of p53, p21, RB and DNA damage. In summary, this thesis identifies TRRAP as a potential oncogene in HCC. I identified a network of genes regulated by TRRAP and its-cofactor KAT5 that promote mitotic progression. Moreover, I demonstrated that disruption of TRRAP/KAT5 and its downstream target gene TOP2A result in senescence of HCC cells independent of p53 status. Taken together, this work suggests that targeting the TRRAP/KAT5 complex and its network of target genes is a potential therapeutic strategy for HCC patients.

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Rubanova, Natalia. "MasterPATH : network analysis of functional genomics screening data". Thesis, Sorbonne Paris Cité, 2018. http://www.theses.fr/2018USPCC109/document.

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Dans ce travail nous avons élaboré une nouvelle méthode de l'analyse de réseau à définir des membres possibles des voies moléculaires qui sont important pour ce phénotype en utilisant la « hit-liste » des expériences « omics » qui travaille dans le réseau intégré (le réseau comprend des interactions protéine-protéine, de transcription, l’acide ribonucléique micro-l’acide ribonucléique messager et celles métaboliques). La méthode tire des sous-réseaux qui sont construit des voies de quatre types les plus courtes (qui ne se composent des interactions protéine-protéine, ayant au minimum une interaction de transcription, ayant au minimum une interaction l’acide ribonucléique micro-l’acide ribonucléique messager, ayant au minimum une interaction métabolique) entre des hit –gènes et des soi-disant « exécuteurs terminaux » - les composants biologiques qui participent à la réalisation du phénotype finale (s’ils sont connus) ou entre les hit-gènes (si « des exécuteurs terminaux » sont inconnus). La méthode calcule la valeur de la centralité de chaque point culminant et de chaque voie dans le sous-réseau comme la quantité des voies les plus courtes trouvées sur la route précédente et passant à travers le point culminant et la voie. L'importance statistique des valeurs de la centralité est estimée en comparaison avec des valeurs de la centralité dans les sous-réseaux construit des voies les plus courtes pour les hit-listes choisi occasionnellement. Il est supposé que les points culminant et les voies avec les valeurs de la centralité statistiquement signifiantes peuvent être examinés comme les membres possibles des voies moléculaires menant à ce phénotype. S’il y a des valeurs expérimentales et la P-valeur pour un grand nombre des points culminant dans le réseau, la méthode fait possible de calculer les valeurs expérimentales pour les voies (comme le moyen des valeurs expérimentales des points culminant sur la route) et les P-valeurs expérimentales (en utilisant la méthode de Fischer et des transpositions multiples).A l'aide de la méthode masterPATH on a analysé les données de la perte de fonction criblage de l’acide ribonucléique micro et l'analyse de transcription de la différenciation terminal musculaire et les données de la perte de fonction criblage du procès de la réparation de l'ADN. On peut trouver le code initial de la méthode si l’on suit le lien https://github.com/daggoo/masterPATH
In this work we developed a new exploratory network analysis method, that works on an integrated network (the network consists of protein-protein, transcriptional, miRNA-mRNA, metabolic interactions) and aims at uncovering potential members of molecular pathways important for a given phenotype using hit list dataset from “omics” experiments. The method extracts subnetwork built from the shortest paths of 4 different types (with only protein-protein interactions, with at least one transcription interaction, with at least one miRNA-mRNA interaction, with at least one metabolic interaction) between hit genes and so called “final implementers” – biological components that are involved in molecular events responsible for final phenotypical realization (if known) or between hit genes (if “final implementers” are not known). The method calculates centrality score for each node and each path in the subnetwork as a number of the shortest paths found in the previous step that pass through the node and the path. Then, the statistical significance of each centrality score is assessed by comparing it with centrality scores in subnetworks built from the shortest paths for randomly sampled hit lists. It is hypothesized that the nodes and the paths with statistically significant centrality score can be considered as putative members of molecular pathways leading to the studied phenotype. In case experimental scores and p-values are available for a large number of nodes in the network, the method can also calculate paths’ experiment-based scores (as an average of the experimental scores of the nodes in the path) and experiment-based p-values (by aggregating p-values of the nodes in the path using Fisher’s combined probability test and permutation approach). The method is illustrated by analyzing the results of miRNA loss-of-function screening and transcriptomic profiling of terminal muscle differentiation and of ‘druggable’ loss-of-function screening of the DNA repair process. The Java source code is available on GitHub page https://github.com/daggoo/masterPATH

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Li, Meng. "Genetic dissection of the exit of pluripotency in mouse embryonic stem cells by CRISPR-based screening". Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/277552.

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The ground state naive pluripotency is established in the epiblast of the blastocyst and can be captured by culturing mouse embryonic stem cells (mESCs) with MEK and GSK3 inhibitors (2i). The transcription network that maintains pluripotency has been extensively studied with the indispensable core factors being Oct4, Sox2 and Nanog, together with other ancillary factors reinforcing the network. However, how this network is dissolved at the onset of differentiation is still not fully understood. To identify genes required for differentiation in an unbiased fashion, I conducted a genome-wide CRISPR-Cas9-mediated screen in Rex1GFPd2 mESCs. This cell line expresses GFP specifically in the naive state and rapidly down-regulate upon differentiation. I differentiated mutagenised mESCs for two days and sorted mutants that kept higher GFP expression. gRNA representation was subsequently analysed by sequencing. I identified 563 and 8 genes whose mutants showed delayed and accelerated differentiation, respectively, at a false discovery rate (FDR) cutoff of 10%. The majority of the previously known genes were identified in my screen, suggesting faithful representation of genes regulating differentiation. Detailed screening result analysis revealed a comprehensive picture of pathways involved in the dissolution of naive pluripotency. Amongst the genes identified are 19 mTORC1 regulators and components of the mTORC2 complex. Deficiency in the TSC and GATOR complexes resulted in mTORC1 upregulation in consistent with previous studies. However, they showed opposite phenotype during ESC differentiation: TSC complex knockout cells showed delayed differentiation, whereas GATOR1 deficiency accelerated differentiation I found that the pattern of GSK3b phosphorylation is highly correlated with differentiation phenotype. I conclude that mTORC1 is involved in pluripotency maintenance and differentiation through cross-talk with the Wnt signalling pathway. My screen has demonstrated the power of CRISPR-Cas9-mediated screen and provided further insights in biological pathways involved in regulating differentiation. It would be interesting to explore the remaining unstudied genes for better understanding of the mechanisms underlying mESC differentiation.

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Kaemena, Daniel Fraser. "CRISPR/Cas9 genome-wide loss of function screening identifies novel regulators of reprogramming to pluripotency". Thesis, University of Edinburgh, 2018. http://hdl.handle.net/1842/31184.

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In 2006, Kazutoshi Takahashi and Shinya Yamanaka demonstrated the ability of four transcription factors; Oct4, Sox2, Klf4 and c-Myc to 'reprogram' differentiated somatic cells to a pluripotent state. This technology holds huge potential in the field of regenerative medicine, but reprogramming also a model system by which to the common regulators of all forced cell identity changes, for example, transdifferentiation. Despite this, the mechanism underlying reprogramming remains poorly understood and the efficiency of induced pluripotent stem cell (iPSC) generation, inefficient. One powerful method for elucidating the gene components influencing a biological process, such as reprogramming, is screening for a phenotype of interest using genome-wide mutant libraries. Historically, large-scale knockout screens have been challenging to perform in diploid mammalian genomes, while other screening technologies such as RNAi can be disadvantaged by variable knockdown of target transcripts and off-target effects. Components of clustered regularly interspaced short palindromic repeats and associated Cas proteins (CRISPR-Cas) prokaryote adaptive immunity systems have recently been adapted to edit genomic sequences at high efficiency in mammalian systems. Furthermore, the application of CRISPR-Cas components to perform proofof- principle genome-wide KO screens has been successfully demonstrated. I have utilised the CRISPR-Cas9 system to perform genome-wide loss-of-function screening in the context of murine iPSC reprogramming, identifying 18 novel inhibitors of reprogramming, in addition to four known inhibitors, Trp53, Cdkn1a, Jun, Dot1l and Gtf2i. Understanding how these novel reprogramming roadblocks function to inhibit the reprogramming process will provide insight into the molecular mechanisms underpinning forced cell identity changes.

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Petrucci, Teresa. "Building a platform for flexible and scalable testing of genetic editors". Doctoral thesis, Università di Siena, 2021. http://hdl.handle.net/11365/1143160.

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Cell-free systems allow to perform in-vitro transcription-translation reactions without requiring living organisms, revolutionising scientific research over the last decade. This allows to easily synthesise a variety of molecular components for genetic editing applications without requiring expensive and time-consuming procedures such as cell culture, animal maintenance etc. In this work, I aimed to develop a high-throughput platform for the rapid, flexible and scalable in-vitro testing of various genetic editors, such as those part of the CRISPR/Cas repertoire. I used the commercially available E. coli cell extract (MyTXTL) in combination with a fully customisable design to generate fluorescent reporters, that allow standardised testing of various CRISPR components against any predesigned target or protospacer adjacent motif (PAM) sequences. In order to increase the scalability of this screening platform, I utilised automated liquid handling technologies (Echo 525) and explored the possibility to introduce a high throughput cloning method (BASIC assembly). I believe that this approach will be highly valuable for the screening of CRISPR components prior than their final application in in-vivo systems, such as humans or animals. These genetic editors could then be used in many biological and artificial systems, such as gene editing, metabolomics and genetic engineering.

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Sczakiel, Henrike Lisa [Verfasser]. "Identifizierung Pathogenese-relevanter Kandidatengene im Hodgkin-Lymphom durch CRISPR/Cas9-basiertes knockout-Screening / Henrike Lisa Sczakiel". Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2021. http://d-nb.info/1228859523/34.

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Lam, Phuong T. "Crispr/cas9-mediated genome editing of human pluripotent stem cells to advance human retina regeneration research". Miami University / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=miami1575372014701457.

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Cresson, Marie. "Study of chikungunya virus entry and host response to infection". Thesis, Lyon, 2019. http://www.theses.fr/2019LYSE1050.

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Les alphavirus sont un groupe de virus enveloppés à ARN simple brin positif retrouvés sur la totalité du globe et responsables de nombreuses maladies humaines et animales. Durant la dernière décennie, une réémergence du virus du chikungunya (CHIKV) a été observée causant de nombreuses épidémies sur tous les continents. Malgré les nombreuses études, les mécanismes moléculaires de réplication du CHIKV et les interactions hôte-virus restent peu caractérisées. L’objectif de mon travail était de mieux comprendre et caractériser l’entrée du virus du chikungunya et les facteurs de l’hôte impliqués dans la réplication chez les mammifères. Plusieurs approches distinctes ont été utilisées dans ce projet. Dans un premier temps, nous avons mis en avant une diminution de l’infection du CHIKV après un traitement avec du fer sous forme de citrate d’ammonium ferrique et nous avons étudié le rôle potentiel dans l’entrée virale de NRAMP2 et TFRC, deux protéines impliquées dans le transport cellulaire du fer et connus comme récepteurs d’entrée de plusieurs virus. D’autre part, nous nous sommes intéressés à deux autres protéines, CD46 et TM9SF2, identifiés à travers un criblage par ARNi réalisé en collaboration, dans le but de déterminer si elles sont utilisées comme facteurs d’entrée par le virus du chikungunya. Dans un dernier axe, nous avons mis en place et réaliser un criblage perte de fonction sur le génome entier en utilisant la technologie CRISPR/Cas9 afin d’identifier des facteurs de l’hôte importants pour l’entrée du CHIKV, sa réplication ou la mort viro-induite. Bien qu’il soit apparu que l’approche utilisée pour le criblage devrait être optimisée, nous avons pu identifier des candidats potentiellement nécessaires pour l’infection par le CHIKV. Ces candidats sont testés individuellement afin de confirmer leur implication dans la biologie du virus
Alphaviruses are a group of enveloped, positive-sense RNA viruses which are distributed almost worldwide and are responsible for a considerable number of human and animal diseases. Among these viruses, the Chikungunya virus (CHIKV) has recently re-emerged and caused several outbreaks on all continents in the past decade. Despite many studies, molecular mechanisms of chikungunya virus replication and virus-host interactions remain poorly understood. The aim of my project was to better understand and characterize the CHIKV entry and the host factors involved during replication steps in mammals. Several different approaches have been used in this work. As a first step, we have demonstrated a decrease of CHIKV infection after iron treatment in form of ferric ammonium citrate and we have studied the potential role in viral entry of NRAMP2 and TFRC, two proteins involved in iron transport and known receptors for other viruses. On the other hand, we have also focused on two proteins, CD46 and TM9SF2, identified through an RNAi screen in collaboration, in order to determine if they are required as entry factors for chikungunya virus. In a last axis, we have set up and carried out a genome-wide loss of function screen with the CRISPR/Cas9 technology in order to identify host factors important for chikungunya virus entry, replication or virus-induced cell death. Although it appears that screen conditions should be optimized, we have identified potential candidates required for CHIKV infection and we are currently testing them

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Mohammad, Jiyan Mageed. "Therapeutic Potential of Piperlongumine for Pancreatic Ductal Adenocarcinoma". Diss., North Dakota State University, 2019. https://hdl.handle.net/10365/31347.

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Pancreatic ductal adenocarcinoma (PDAC) is among the most lethal malignancies because it is often diagnosed at a late disease stage and has a poor response rate to currently available treatments. Therefore, it is critical to develop new therapeutic approaches that will enhance the efficacy and reduce the toxicity of currently used therapies. Here we aimed to evaluate the therapeutic potential and mechanisms of action for piperlongumine (PL), an alkaloid from long pepper, in PDAC models. We postulated that PL causes PDAC cell death through oxidative stress and complements the therapeutic efficacy of chemotherapeutic agents in PDAC cells. First, we determined that PL is one of the most abundant alkaloids with antitumor properties in the long pepper plant. We also showed PL in combination with gemcitabine, a chemotherapy agent used to treat advanced pancreatic cancer, reduced tumor weight and volume compared to vehicle-control and individual treatments. Further, biochemical analysis, including RNA sequencing and immunohistochemistry, suggested that the antitumor activity of PL was associated with decreased cell proliferation, induction of cell cycle arrest, and oxidative stress-induced cell death. Moreover, we identified that c-Jun N-terminal kinase (JNK) inhibition blocks PL-induced cell death, translocation of Nrf2, and transcriptional activation of HMOX1 in PDAC. Finally, high-throughput drug and CRISPR screenings identified potential targets that could be used in combination with PL to treat PDAC cells. Collectively, our data suggests that cell cycle regulators in combination with PL might be an effective approach to combat pancreatic cancer.
NIH

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Livros sobre o assunto "CRISPRko Screening"

Barilan, Y. Michael, Margherita Brusa e Aaron Ciechanover, eds. Can precision medicine be personal; Can personalized medicine be precise? Oxford University Press, 2022. http://dx.doi.org/10.1093/oso/9780198863465.001.0001.

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In the medicine of the fifty years, the prevailing paradigms were the ‘biopsychosocial model’ and ‘evidence-based medicine’. The currently emerging vision is ‘personalized/precision medicine’. The two interchangeable names—personalized medicine and precision medicine—raise fundamental questions. Is increased precision an improvement in the personal aspects of care or does it risk an erosion of privacy? Do ‘precise’ and ‘personalized’ approaches marginalize public health? What are the roles of culture and society in the process of personalization? How can personalized medicine’s focus on the differences among individuals contribute to the global enterprise of health? In this project, scientists who are leading the revolution of personalized medicine are engaged with clinicians, ethicists, and experts in sociology of medicine and medical law in the search for a common language, elucidating and discussing the moral and social dimensions of personalized/precision medicine. The result is diverse layers of critical analysis and insights. The book contains eighteen chapters. The opening chapters map the horizon of the discourse, articulating the vision and practice of personalized medicine in the contexts of the history of ideas, philosophy of science, and global health. The subsequent chapters explicate specific topics: genetic newborn screening, rare diseases, disorders of consciousness, gender, the clinical encounter, public health, and CRISPR. The concluding chapters offer critical reflections by academic science and technology studies, and by religious traditions. The book concludes by presenting an up-to-date overview from the perspective of research and development.

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Capítulos de livros sobre o assunto "CRISPRko Screening"

Henriksson, Johan. "CRISPR Screening in Single Cells". In Methods in Molecular Biology, 395–406. New York, NY: Springer New York, 2019. http://dx.doi.org/10.1007/978-1-4939-9240-9_23.

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Nishiga, Masataka, Lei S. Qi e Joseph C. Wu. "CRISPRi/a Screening with Human iPSCs". In Methods in Molecular Biology, 261–81. New York, NY: Springer US, 2021. http://dx.doi.org/10.1007/978-1-0716-1484-6_23.

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Port, Fillip, e Michael Boutros. "Tissue-Specific CRISPR-Cas9 Screening in Drosophila". In Methods in Molecular Biology, 157–76. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2541-5_7.

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AbstractOver the last century research in Drosophila has resulted in many fundamental contributions to our understanding of the biology of multicellular organisms. Many of these breakthroughs have been based on the identification of novel gene functions in large-scale genetic screens. However, conventional forward-genetic screens have been limited by the random nature of mutagenesis and difficulties in mapping causal mutations, while reverse-genetic RNAi screens suffer from incomplete knockdown of gene expression. Recently developed large-scale CRISPR-Cas9 libraries promise to address these limitations by allowing the induction of targeted mutations in genes with spatial and temporal control. Here, we provide a guide for tissue-specific CRISPR screening in Drosophila, including the characterization of Gal4 UAS-Cas9 lines, selection of sgRNA libraries, and various quality control measures. We also discuss confounding factors that can give rise to false-positive and false-negative results in such experiments and suggest strategies on how to detect and avoid them. Conditional CRISPR screening represents an exciting new approach for functional genomics in vivo and is set to further expand our knowledge of the molecular underpinning of development, homeostasis, and disease.

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DeLuca, Sophia, e Nenad Bursac. "CRISPR Library Screening in Cultured Cardiomyocytes". In Methods in Molecular Biology, 1–13. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2261-2_1.

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Carstens, Carsten P., Katherine A. Felts e Sarah E. Johns. "Construction of CRISPR Libraries for Functional Screening". In Synthetic Biology, 139–50. New York, NY: Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-7795-6_7.

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Webster, Daniel E., Sandrine Roulland e James D. Phelan. "Protocols for CRISPR-Cas9 Screening in Lymphoma Cell Lines". In Methods in Molecular Biology, 337–50. New York, NY: Springer New York, 2019. http://dx.doi.org/10.1007/978-1-4939-9151-8_16.

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Haney, Steven A. "High-Content Screening Approaches That Minimize Confounding Factors in RNAi, CRISPR, and Small Molecule Screening". In Methods in Molecular Biology, 113–30. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7357-6_8.

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Klann, Tyler S., Gregory E. Crawford, Timothy E. Reddy e Charles A. Gersbach. "Screening Regulatory Element Function with CRISPR/Cas9-based Epigenome Editing". In Methods in Molecular Biology, 447–80. New York, NY: Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-7774-1_25.

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Carlini, Valentina, Kristjan H. Gretarsson e Jamie A. Hackett. "Genome-Scale CRISPR Screening for Regulators of Cell Fate Transitions". In Methods in Molecular Biology, 91–108. New York, NY: Springer US, 2020. http://dx.doi.org/10.1007/978-1-0716-0958-3_7.

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Shang, Wanjing, Fei Wang, Qi Zhu, Liangyu Wang e Haopeng Wang. "CRISPR/Cas9-Based Genetic Screening to Study T-Cell Function". In Methods in Molecular Biology, 59–70. New York, NY: Springer US, 2020. http://dx.doi.org/10.1007/978-1-0716-0266-9_5.

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Trabalhos de conferências sobre o assunto "CRISPRko Screening"

le Sage, Carlos, Prince Panckier e Bendict CS Cross. "Abstract A186: Dual CRISPRi and CRISPRa screening reveals phenotypic switches in response to BRAF inhibition". In Abstracts: AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; October 26-30, 2017; Philadelphia, PA. American Association for Cancer Research, 2018. http://dx.doi.org/10.1158/1535-7163.targ-17-a186.

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Cross, Benedict CS, Steffen Lawo, Tim ME Scales, Caroline Archer, Jessica Hunt, Alessandro Riccombeni, Leigh Brody et al. "Abstract B163: Genetic screening with CRISPR-Cas9: Proof and principles". In Abstracts: AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; November 5-9, 2015; Boston, MA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1535-7163.targ-15-b163.

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Tedesco, Donato, Paul Diehl, Mikhail Makhanov, Sylvain Baron, Dmitry Suchkov e Alex Chenchik. "Abstract C161: CRISPR/Cas9 genome-wide gRNA library screening platform". In Abstracts: AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; November 5-9, 2015; Boston, MA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1535-7163.targ-15-c161.

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Tan, Jenille M., e Scott E. Martin. "Abstract 73: Exploring arrayed synthetic CRISPR for functional genomic screening". In Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LA. American Association for Cancer Research, 2016. http://dx.doi.org/10.1158/1538-7445.am2016-73.

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Li, Wanji. "Development and Application of CRISPR-Mediated Genetic Screening in Oncology". In ICBBE '20: 2020 7th International Conference on Biomedical and Bioinformatics Engineering. New York, NY, USA: ACM, 2020. http://dx.doi.org/10.1145/3444884.3444909.

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Wei, Yiliang, e Christopher Vakoc. "Abstract 5161: Probing leukemia vulnerabilitiesin vivousing domain-focused CRISPR screening". In Proceedings: AACR Annual Meeting 2019; March 29-April 3, 2019; Atlanta, GA. American Association for Cancer Research, 2019. http://dx.doi.org/10.1158/1538-7445.sabcs18-5161.

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Grassian, Alexandra R., Julian Fowler, Igor Feldman, Thomas Riera, Darren Harvey, Allison E. Drew, Richard Chesworth et al. "Abstract B78: CRISPR pooled screening identifies differential dependencies on epigenetic pathways". In Abstracts: AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; November 5-9, 2015; Boston, MA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1535-7163.targ-15-b78.

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Vakoc, Christopher. "Abstract IA07: Cancer drug target identification using domain-focused CRISPR screening". In Abstracts: Advances in Sarcomas: From Basic Science to Clinical Translation; May 16-19, 2017; Philadelphia, PA. American Association for Cancer Research, 2018. http://dx.doi.org/10.1158/1557-3265.sarcomas17-ia07.

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Behan, F., F. Iorio, E. Stronach, C. Beaver, R. Moita Santos, J. Saez-Rodriguez, K. Yusa e M. Garnett. "SPOT-012 Large-scale CRISPR screening to identify actionable cancer drug targets". In Abstracts of the 25th Biennial Congress of the European Association for Cancer Research, Amsterdam, The Netherlands, 30 June – 3 July 2018. BMJ Publishing Group Ltd, 2018. http://dx.doi.org/10.1136/esmoopen-2018-eacr25.45.

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