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Nature Biotechnology

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https://www.readbyqxmd.com/read/29786095/high-throughput-creation-and-functional-profiling-of-dna-sequence-variant-libraries-using-crispr-cas9-in-yeast
#1
Xiaoge Guo, Alejandro Chavez, Angela Tung, Yingleong Chan, Christian Kaas, Yi Yin, Ryan Cecchi, Santiago Lopez Garnier, Eric D Kelsic, Max Schubert, James E DiCarlo, James J Collins, George M Church
Construction and characterization of large genetic variant libraries is essential for understanding genome function, but remains challenging. Here, we introduce a Cas9-based approach for generating pools of mutants with defined genetic alterations (deletions, substitutions, and insertions) with an efficiency of 80-100% in yeast, along with methods for tracking their fitness en masse. We demonstrate the utility of our approach by characterizing the DNA helicase SGS1 with small tiling deletion mutants that span the length of the protein and a series of point mutations against highly conserved residues in the protein...
May 21, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29786094/a-protein-activity-assay-to-measure-global-transcription-factor-activity-reveals-determinants-of-chromatin-accessibility
#2
Bei Wei, Arttu Jolma, Biswajyoti Sahu, Lukas M Orre, Fan Zhong, Fangjie Zhu, Teemu Kivioja, Inderpreet Sur, Janne Lehtiö, Minna Taipale, Jussi Taipale
No existing method to characterize transcription factor (TF) binding to DNA allows genome-wide measurement of all TF-binding activity in cells. Here we present a massively parallel protein activity assay, active TF identification (ATI), that measures the DNA-binding activity of all TFs in cell or tissue extracts. ATI is based on electrophoretic separation of protein-bound DNA sequences from a highly complex DNA library and subsequent mass-spectrometric identification of the DNA-bound proteins. We applied ATI to four mouse tissues and mouse embryonic stem cells and found that, in a given tissue or cell type, a small set of TFs, which bound to only ∼10 distinct motifs, displayed strong DNA-binding activity...
May 21, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29786096/reversal-of-sirna-mediated-gene-silencing-in-vivo
#3
Ivan Zlatev, Adam Castoreno, Christopher R Brown, June Qin, Scott Waldron, Mark K Schlegel, Rohan Degaonkar, Svetlana Shulga-Morskaya, Huilei Xu, Swati Gupta, Shigeo Matsuda, Akin Akinc, Kallanthottathil G Rajeev, Muthiah Manoharan, Martin A Maier, Vasant Jadhav
We report rapid, potent reversal of GalNAc-siRNA-mediated RNA interference (RNAi) activity in vivo with short, synthetic, high-affinity oligonucleotides complementary to the siRNA guide strand. We found that 9-mers with five locked nucleic acids (LNAs) have the highest potency across several targets. Our modular, sequence-specific approach, named REVERSIR, may enhance the therapeutic profile of any long-acting GalNAc-siRNA (short interfering RNA) conjugate by enabling control of RNAi pharmacology.
May 14, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734295/genome-scale-engineering-of-saccharomyces-cerevisiae-with-single-nucleotide-precision
#4
Zehua Bao, Mohammad HamediRad, Pu Xue, Han Xiao, Ipek Tasan, Ran Chao, Jing Liang, Huimin Zhao
We developed a CRISPR-Cas9- and homology-directed-repair-assisted genome-scale engineering method named CHAnGE that can rapidly output tens of thousands of specific genetic variants in yeast. More than 98% of target sequences were efficiently edited with an average frequency of 82%. We validate the single-nucleotide resolution genome-editing capability of this technology by creating a genome-wide gene disruption collection and apply our method to improve tolerance to growth inhibitors.
May 7, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734294/multiplexed-precision-genome-editing-with-trackable-genomic-barcodes-in-yeast
#5
Kevin R Roy, Justin D Smith, Sibylle C Vonesch, Gen Lin, Chelsea Szu Tu, Alex R Lederer, Angela Chu, Sundari Suresh, Michelle Nguyen, Joe Horecka, Ashutosh Tripathi, Wallace T Burnett, Maddison A Morgan, Julia Schulz, Kevin M Orsley, Wu Wei, Raeka S Aiyar, Ronald W Davis, Vytas A Bankaitis, James E Haber, Marc L Salit, Robert P St Onge, Lars M Steinmetz
Our understanding of how genotype controls phenotype is limited by the scale at which we can precisely alter the genome and assess the phenotypic consequences of each perturbation. Here we describe a CRISPR-Cas9-based method for multiplexed accurate genome editing with short, trackable, integrated cellular barcodes (MAGESTIC) in Saccharomyces cerevisiae. MAGESTIC uses array-synthesized guide-donor oligos for plasmid-based high-throughput editing and features genomic barcode integration to prevent plasmid barcode loss and to enable robust phenotyping...
May 7, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734293/secure-genome-wide-association-analysis-using-multiparty-computation
#6
Hyunghoon Cho, David J Wu, Bonnie Berger
Most sequenced genomes are currently stored in strict access-controlled repositories. Free access to these data could improve the power of genome-wide association studies (GWAS) to identify disease-causing genetic variants and aid the discovery of new drug targets. However, concerns over genetic data privacy may deter individuals from contributing their genomes to scientific studies and could prevent researchers from sharing data with the scientific community. Although cryptographic techniques for secure data analysis exist, none scales to computationally intensive analyses, such as GWAS...
May 7, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29658944/an-in-vivo-model-of-functional-and-vascularized-human-brain-organoids
#7
Abed AlFatah Mansour, J Tiago Gonçalves, Cooper W Bloyd, Hao Li, Sarah Fernandes, Daphne Quang, Stephen Johnston, Sarah L Parylak, Xin Jin, Fred H Gage
Differentiation of human pluripotent stem cells to small brain-like structures known as brain organoids offers an unprecedented opportunity to model human brain development and disease. To provide a vascularized and functional in vivo model of brain organoids, we established a method for transplanting human brain organoids into the adult mouse brain. Organoid grafts showed progressive neuronal differentiation and maturation, gliogenesis, integration of microglia, and growth of axons to multiple regions of the host brain...
April 16, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29658943/deep-learning-massively-accelerates-super-resolution-localization-microscopy
#8
Wei Ouyang, Andrey Aristov, Mickaël Lelek, Xian Hao, Christophe Zimmer
The speed of super-resolution microscopy methods based on single-molecule localization, for example, PALM and STORM, is limited by the need to record many thousands of frames with a small number of observed molecules in each. Here, we present ANNA-PALM, a computational strategy that uses artificial neural networks to reconstruct super-resolution views from sparse, rapidly acquired localization images and/or widefield images. Simulations and experimental imaging of microtubules, nuclear pores, and mitochondria show that high-quality, super-resolution images can be reconstructed from up to two orders of magnitude fewer frames than usually needed, without compromising spatial resolution...
April 16, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29644997/highly-scalable-generation-of-dna-methylation-profiles-in-single-cells
#9
Ryan M Mulqueen, Dmitry Pokholok, Steven J Norberg, Kristof A Torkenczy, Andrew J Fields, Duanchen Sun, John R Sinnamon, Jay Shendure, Cole Trapnell, Brian J O'Roak, Zheng Xia, Frank J Steemers, Andrew C Adey
We present a highly scalable assay for whole-genome methylation profiling of single cells. We use our approach, single-cell combinatorial indexing for methylation analysis (sci-MET), to produce 3,282 single-cell bisulfite sequencing libraries and achieve read alignment rates of 68 ± 8%. We apply sci-MET to discriminate the cellular identity of a mixture of three human cell lines and to identify excitatory and inhibitory neuronal populations from mouse cortical tissue.
April 9, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29644996/simultaneous-lineage-tracing-and-cell-type-identification-using-crispr-cas9-induced-genetic-scars
#10
Bastiaan Spanjaard, Bo Hu, Nina Mitic, Pedro Olivares-Chauvet, Sharan Janjuha, Nikolay Ninov, Jan Philipp Junker
A key goal of developmental biology is to understand how a single cell is transformed into a full-grown organism comprising many different cell types. Single-cell RNA-sequencing (scRNA-seq) is commonly used to identify cell types in a tissue or organ. However, organizing the resulting taxonomy of cell types into lineage trees to understand the developmental origin of cells remains challenging. Here we present LINNAEUS (lineage tracing by nuclease-activated editing of ubiquitous sequences)-a strategy for simultaneous lineage tracing and transcriptome profiling in thousands of single cells...
April 9, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29608177/batch-effects-in-single-cell-rna-sequencing-data-are-corrected-by-matching-mutual-nearest-neighbors
#11
Laleh Haghverdi, Aaron T L Lun, Michael D Morgan, John C Marioni
Large-scale single-cell RNA sequencing (scRNA-seq) data sets that are produced in different laboratories and at different times contain batch effects that may compromise the integration and interpretation of the data. Existing scRNA-seq analysis methods incorrectly assume that the composition of cell populations is either known or identical across batches. We present a strategy for batch correction based on the detection of mutual nearest neighbors (MNNs) in the high-dimensional expression space. Our approach does not rely on predefined or equal population compositions across batches; instead, it requires only that a subset of the population be shared between batches...
April 2, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29608178/simultaneous-single-cell-profiling-of-lineages-and-cell-types-in-the-vertebrate-brain
#12
Bushra Raj, Daniel E Wagner, Aaron McKenna, Shristi Pandey, Allon M Klein, Jay Shendure, James A Gagnon, Alexander F Schier
The lineage relationships among the hundreds of cell types generated during development are difficult to reconstruct. A recent method, GESTALT, used CRISPR-Cas9 barcode editing for large-scale lineage tracing, but was restricted to early development and did not identify cell types. Here we present scGESTALT, which combines the lineage recording capabilities of GESTALT with cell-type identification by single-cell RNA sequencing. The method relies on an inducible system that enables barcodes to be edited at multiple time points, capturing lineage information from later stages of development...
March 28, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29644998/fast-long-term-super-resolution-imaging-with-hessian-structured-illumination-microscopy
#13
Xiaoshuai Huang, Junchao Fan, Liuju Li, Haosen Liu, Runlong Wu, Yi Wu, Lisi Wei, Heng Mao, Amit Lal, Peng Xi, Liqiang Tang, Yunfeng Zhang, Yanmei Liu, Shan Tan, Liangyi Chen
To increase the temporal resolution and maximal imaging time of super-resolution (SR) microscopy, we have developed a deconvolution algorithm for structured illumination microscopy based on Hessian matrixes (Hessian-SIM). It uses the continuity of biological structures in multiple dimensions as a priori knowledge to guide image reconstruction and attains artifact-minimized SR images with less than 10% of the photon dose used by conventional SIM while substantially outperforming current algorithms at low signal intensities...
June 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29608179/integrating-single-cell-transcriptomic-data-across-different-conditions-technologies-and-species
#14
Andrew Butler, Paul Hoffman, Peter Smibert, Efthymia Papalexi, Rahul Satija
Computational single-cell RNA-seq (scRNA-seq) methods have been successfully applied to experiments representing a single condition, technology, or species to discover and define cellular phenotypes. However, identifying subpopulations of cells that are present across multiple data sets remains challenging. Here, we introduce an analytical strategy for integrating scRNA-seq data sets based on common sources of variation, enabling the identification of shared populations across data sets and downstream comparative analysis...
June 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734317/alternative-models-for-sharing-confidential-biomedical-data
#15
Justin Guinney, Julio Saez-Rodriguez
No abstract text is available yet for this article.
May 9, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734316/a-rich-resource
#16
Ahmet-Hamdi Cavusoglu, Elizabeth Beerman, Orin Herskowitz
No abstract text is available yet for this article.
May 9, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734315/all-the-skin-that-s-fit-to-print
#17
Irene Jarchum
No abstract text is available yet for this article.
May 9, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734314/boosting-the-power-of-single-cell-analysis
#18
Lu Wen, Fuchou Tang
No abstract text is available yet for this article.
May 9, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734313/around-the-world-in-a-month
#19
(no author information available yet)
No abstract text is available yet for this article.
May 9, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29734312/first-quarter-biotech-job-picture
#20
Michael Francisco
No abstract text is available yet for this article.
May 9, 2018: Nature Biotechnology
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