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Improved design and analysis of CRISPR knockout screens

Chen-Hao Chen, Tengfei Xiao, Han Xu, Peng Jiang, Clifford A Meyer, Wei Li, Myles Brown, X Shirley Liu, Jonathan Wren
2018 Bioinformatics  
Collectively, our analysis motivated the design of a new genome-wide CRISPR/Cas9 screen library and uncovered some intriguing properties of the CRISPR-Cas9 system.  ...  Custom-designed screens confirmed our findings and further revealed that 19nt sgRNAs consistently gave the best signal-to-noise ratio.  ...  Collectively, our study provided novel insights the properties of CRISPR and the design of both high-and low throughput CRISPR experiments.  ... 
doi:10.1093/bioinformatics/bty450 pmid:29868757 fatcat:2buh4gcpqfhlzaxrzhuh5xhicm

To Discover the Efficient and Novel Drug Targets in Human Cancers Using CRISPR/Cas Screening and Databases

Iichiroh Onishi, Kouhei Yamamoto, Yuko Kinowaki, Masanobu Kitagawa, Morito Kurata
2021 International Journal of Molecular Sciences  
However, it is easier to efficiently search for cancer targets with their own screenings by effectively using databases of CRISPR screenings, such as Depmap portal, PICKLES (Pooled In-Vitro CRISPR Knockout  ...  This review will suggest recent optimal CRISPR libraries and effective databases for Novel Approaches in the Discovery and Design of Targeted Therapies.  ...  Conflicts of Interest: The authors declare no conflict of interest.  ... 
doi:10.3390/ijms222212322 pmid:34830205 pmcid:PMC8622676 fatcat:vq7yzl2y3zfm3oghi7yqk272g4

CRISPR screen in cancer: status quo and future perspectives

Chenglong He, Siqi Han, Yue Chang, Meijuan Wu, Yulu Zhao, Cheng Chen, Xiaoyuan Chu
2021 American Journal of Cancer Research  
and future perspectives for CRISPR screen.  ...  Key parameters of CRISPR screen are constantly being optimized to achieve higher targeting efficiency and lower off-target effect.  ...  Acknowledgements This work was supported by grants from National Natural Science Foundation of China (81872042 and 81972333), Foundation of Jiangsu Province (BK20181238) and Jiangsu  ... 
pmid:33948344 pmcid:PMC8085856 fatcat:a57jlropgvg4zgbgwd4gzc3g34

Research Techniques Made Simple: CRISPR Genetic Screens

Auke B.C. Otten, Bryan K. Sun
2020 Journal of Investigative Dermatology  
Here, we discuss major considerations in the design, execution, and analysis of CRISPR screens.  ...  We focus on CRISPR knockout screens but also review adaptations to the CRISPR/Cas system that highlight the versatility of the system to make other types of experimental genetic changes as well.  ...  Alan O'Neill and Dr. Prashant Mali for their critical evaluation and improvements to this manuscript.  ... 
doi:10.1016/j.jid.2020.01.018 pmid:32200874 pmcid:PMC8525197 fatcat:zjt4vhvuznfxrnq7tcajpgjswq

High-performance CRISPR-Cas12a genome editing for combinatorial genetic screening

Rodrigo A. Gier, Krista A. Budinich, Niklaus H. Evitt, Zhendong Cao, Elizabeth S. Freilich, Qingzhou Chen, Jun Qi, Yemin Lan, Rahul M. Kohli, Junwei Shi
2020 Nature Communications  
We demonstrate the performance of our optimized AsCas12a (opAsCas12a) through double knockout screening against epigenetic regulators.  ...  Here, we present a simple and robust CRISPR-Cas12a-based approach for combinatorial genetic screening in cancer cells.  ...  J.S. acknowledges support from the Epigenetics Institute at the University of Pennsylvania pilot award and the Linda Pechenik Montague Investigator Award.  ... 
doi:10.1038/s41467-020-17209-1 pmid:32661245 fatcat:tly5yquc25cqpnsgqn2yjkpktm

CRISPR-cas9: a powerful tool towards precision medicine in cancer treatment

Hui Xing, Ling-Hua Meng
2019 Acta Pharmacologica Sinica  
In this review, we describe the progress of CRISPR-cas9-based unbiased screening in precision medicine including identification of new drug targets, biomarkers and elucidation of mechanisms leading to  ...  CRISPR-cas9 system is a powerful tool for gene editing and CRISPR-based high-throughput screening has been widely applied especially in searching for tumor-driven or synergistic lethal genes aiming to  ...  The improvement of the delivery system and CRISPR-cas9 efficiency will be critical for improved screening in vivo.  ... 
doi:10.1038/s41401-019-0322-9 pmid:31792341 pmcid:PMC7468325 fatcat:p7h36uwpzvc6tpspdidrszib2i

Sequence determinants of improved CRISPR sgRNA design

Han Xu, Tengfei Xiao, Chen-Hao Chen, Wei Li, Clifford A. Meyer, Qiu Wu, Di Wu, Le Cong, Feng Zhang, Jun S. Liu, Myles Brown, X. Shirley Liu
2015 Genome Research  
These results facilitate the genome-wide design of improved sgRNA for both knockout and CRISPRi/a studies.  ...  Leveraging the information from multiple designs, we derived a new sequence model for predicting sgRNA efficiency in CRISPR/Cas9 knockout experiments.  ...  Carl Novina, Tim Wang, David Sabatini, Eric Lander, Ophir Shalem, and Neville Sanjana for providing the data sets and helpful discussions.  ... 
doi:10.1101/gr.191452.115 pmid:26063738 pmcid:PMC4509999 fatcat:epd2wtpa7ba23n7jtj46shp5nq

Pooled CRISPR screening at high sensitivity with an empirically designed sgRNA library [article]

Luisa Henkel, Benedikt Rauscher, Barbara Schmitt, Jan Winter, Michael Boutros
2020 bioRxiv   pre-print
Given their broad utility in functionally annotating genomes, the experimental design of genome-scale CRISPR screens can vary greatly and criteria for optimal experimental implementation and library composition  ...  We further systematically analyzed published CRISPR screens in human cells to identify single-guide (sg)RNAs with consistent high on-target and low off-target activity.  ...  In recent years, the design of sgRNAs and CRISPR libraries was improved based on design rules derived from comparing the nucleotide composition of active and non-active guides (Chen et al. 2018; Tzelepis  ... 
doi:10.1101/2020.04.25.061135 fatcat:lqgkqtuoizayhdzaezs2sef2ie

Genome-scale CRISPR screening at high sensitivity with an empirically designed sgRNA library

Luisa Henkel, Benedikt Rauscher, Barbara Schmitt, Jan Winter, Michael Boutros
2020 BMC Biology  
Given their broad utility in functionally annotating genomes, the application and interpretation of genome-scale CRISPR screens would greatly benefit from consistent and optimal design criteria.  ...  However, screening designs vary greatly and criteria for optimal experimental implementation and library composition are still emerging.  ...  Acknowledgements We thank Fillip Port for help with the design of the cloning strategy and Josephine Bageritz for help with FACS analysis.  ... 
doi:10.1186/s12915-020-00905-1 pmid:33228647 fatcat:abjcifxu3bg2rkpzqiduk6b72m

CRISPR/Cas9‐mediated knockout of an oil palm defense‐related gene to the pathogenic fungus Ganoderma boninense

Asmini Budiani, Imam Bagus Nugroho, Dini Astika Sari, Inez Palupi, Riza Arief Putranto
2020 Indonesian Journal of Biotechnology  
The transformed OPCs were regenerated and screened in DF (de Fossard) media containing hygromycin. The working concentration of hygromycin was successfully optimized for selection at 20 ppm.  ...  Moreover, electrophoresis analysis of PCR products of cDNA as the template from transformed OPC clones showed several samples with faint or undetected bands.  ...  Acknowledgments The authors thank Marini, Elizabeth Monica Sitorus, and Nazhirotul Ilmiyah for the assistance. This research was funded by the BPDPKS (Indonesian Oil Palm Research Funding Agency).  ... 
doi:10.22146/ijbiotech.52170 fatcat:he2rqhdzbjcofjrcm5bn7o7m54

RAB18 is a key regulator of GalNAc conjugated siRNA induced silencing in Hep3B cells [article]

Jiamiao Lu, Elissa Swearingen, Miki Hardy, Patrick Collins, Bin Wu, Eric Yuan, Daniel Lu, Chi-Ming Kevin Li, Songli Wang, Michael Ollmann
2022 bioRxiv   pre-print
Follow-up studies indicate that knockout of one gene in particular, RAB18, improved siRNA efficacy.  ...  To reveal the cellular factors limiting delivery of siRNA therapeutics, we performed a pooled, genome wide knockout screen based on delivery of GalNAc conjugated siRNA targeting the HPRT1 gene in the human  ...  ACKNOWLEDGEMENTS We thank Stephen Wong and Oliver Homann for providing input for NGS data analysis. We also thank Karen Siegler for providing 7E11 antibody.  ... 
doi:10.1101/2022.01.07.475411 fatcat:pmtrcfpxwfeexpl5mg6gucqw5e

MAGeCK enables robust identification of essential genes from genome-scale CRISPR/Cas9 knockout screens

Wei Li, Han Xu, Tengfei Xiao, Le Cong, Michael I Love, Feng Zhang, Rafael A Irizarry, Jun S Liu, Myles Brown, X Liu
2014 Genome Biology  
We propose the Model-based Analysis of Genome-wide CRISPR/Cas9 Knockout (MAGeCK) method for prioritizing single-guide RNAs, genes and pathways in genome-scale CRISPR/Cas9 knockout screens.  ...  MAGeCK demonstrates better performance compared with existing methods, identifies both positively and negatively selected genes simultaneously, and reports robust results across different experimental  ...  Acknowledgements The authors would like to thank Drs Tim Wang, Eric Lander, David Sabatini, Ophir Shalem, Neville Sanjana, Graham McVicker and Clifford Meyer for providing the datasets and helpful discussions  ... 
doi:10.1186/preaccept-1316450832143458 pmid:25476604 pmcid:PMC4290824 fatcat:udoepuphlzb6lcgsgqmdwdu5mu

GEMINI: a variational Bayesian approach to identify genetic interactions from combinatorial CRISPR screens

Mahdi Zamanighomi, Sidharth S. Jain, Takahiro Ito, Debjani Pal, Timothy P. Daley, William R. Sellers
2019 Genome Biology  
The improved accuracy and scalability of GEMINI enables the systematic analysis of combinatorial CRISPR knockout screens, regardless of design and dimension.  ...  We develop a variational Bayes approach (GEMINI) that jointly analyzes all samples and reagents to identify genetic interactions in pairwise knockout screens.  ...  James McFarland, and Dr. Josh Dempster, as well as two anonymous reviewers, for helpful suggestions and feedback. Review history The review history is included as Additional file 3.  ... 
doi:10.1186/s13059-019-1745-9 fatcat:ke3zfzkguzhyllrdhe2r6vgipa

Evaluation and Design of Genome-Wide CRISPR/SpCas9 Knockout Screens

Traver Hart, Amy Hin Yan Tong, Katie Chan, Jolanda Van Leeuwen, Ashwin Seetharaman, Michael Aregger, Megha Chandrashekhar, Nicole Hustedt, Sahil Seth, Avery Noonan, Andrea Habsid, Olga Sizova (+20 others)
2017 G3: Genes, Genomes, Genetics  
We use this expanded set of reference core essential genes, CEG2, plus empirical data from six CRISPR knockout screens to guide the design of a sequence-optimized gRNA library, the Toronto KnockOut version  ...  designs and experimental protocols.  ...  genetic screens CRISPR/Cas9 core essential genes cancer cell lines The generation of gene knockouts is a cornerstone of functional genomics research.  ... 
doi:10.1534/g3.117.041277 pmid:28655737 pmcid:PMC5555476 fatcat:jwjzzawrirh6lhzxecxouyv6ce

RNAi screening comes of age: improved techniques and complementary approaches

Stephanie E. Mohr, Jennifer A. Smith, Caroline E. Shamu, Ralph A. Neumüller, Norbert Perrimon
2014 Nature reviews. Molecular cell biology  
Considerable progress has been made to improve the quality of RNAi screen data through the development of new experimental and bioinformatics approaches.  ...  in screen data quality and follow-up experiments, thus promoting our understanding of gene function and gene regulatory networks.  ...  Acknowledgements The Drosophila RNAi Screening Center is supported by US National Institutes of Health (NIH) NIGMS R01 GM067761 (N.P.).  ... 
doi:10.1038/nrm3860 pmid:25145850 pmcid:PMC4204798 fatcat:rjkc3uyi3rc3vdn7rkgvpnjy6q
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