显示样式:     当前期刊: Nature Methods    加入关注       排序: 导出
我的关注
我的收藏
您暂时未登录!
登录
  • Real-time 3D single-molecule localization using experimental point spread functions
    Nat. Methods (IF 25.062) Pub Date : 2018-04-09
    Yiming Li, Markus Mund, Philipp Hoess, Joran Deschamps, Ulf Matti, Bianca Nijmeijer, Vilma Jimenez Sabinina, Jan Ellenberg, Ingmar Schoen, Jonas Ries

    We present a real-time fitter for 3D single-molecule localization microscopy using experimental point spread functions (PSFs) that achieves minimal uncertainty in 3D on any microscope and is compatible with any PSF engineering approach. We used this method to image cellular structures and attained unprecedented image quality for astigmatic PSFs. The fitter compensates for most optical aberrations and makes accurate 3D super-resolution microscopy broadly accessible, even on standard microscopes without dedicated 3D optics.

    更新日期:2018-04-09
  • FateID infers cell fate bias in multipotent progenitors from single-cell RNA-seq data
    Nat. Methods (IF 25.062) Pub Date : 2018-04-09
    Josip S Herman, Sagar, Dominic Grün

    To understand stem cell differentiation along multiple lineages, it is necessary to resolve heterogeneous cellular states and the ancestral relationships between them. We developed a robotic miniaturized CEL-Seq2 implementation to carry out deep single-cell RNA-seq of ∼2,000 mouse hematopoietic progenitors enriched for lymphoid lineages, and used an improved clustering algorithm, RaceID3, to identify cell types. To resolve subtle transcriptome differences indicative of lineage biases, we developed FateID, an iterative supervised learning algorithm for the probabilistic quantification of cell fate bias in progenitor populations. Here we used FateID to delineate domains of fate bias and enable the derivation of high-resolution differentiation trajectories, thereby revealing a common progenitor population of B cells and plasmacytoid dendritic cells, which we validated by in vitro differentiation assays. We expect that FateID will improve understanding of the process of cell fate choice in complex multi-lineage differentiation systems.

    更新日期:2018-04-09
  • How to pull the blanket off dormant cancer cells
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03
    Vivien Marx

    How to pull the blanket off dormant cancer cells How to pull the blanket off dormant cancer cells, Published online: 03 April 2018; doi:10.1038/nmeth.4640 When asleep, cancer cells can evade chemo. When they wake up, they can cause cancer recurrence. By deciphering dormancy cues, labs explore how to break this cycle.

    更新日期:2018-04-04
  • Points of Significance: Statistics versus machine learning
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03
    Danilo Bzdok, Naomi Altman, Martin Krzywinski

    Points of Significance: Statistics versus machine learning Points of Significance: Statistics versus machine learning, Published online: 03 April 2018; doi:10.1038/nmeth.4642 Statistics draws population inferences from a sample, and machine learning finds generalizable predictive patterns.

    更新日期:2018-04-04
  • Genomics: Single-cell hat trick
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03

    Genomics: Single-cell hat trick Genomics: Single-cell hat trick, Published online: 03 April 2018; doi:10.1038/nmeth.4646 Genomics: Single-cell hat trick

    更新日期:2018-04-04
  • Genomics: Off-targets in epigenome editing
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03

    Genomics: Off-targets in epigenome editing Genomics: Off-targets in epigenome editing, Published online: 03 April 2018; doi:10.1038/nmeth.4650 Genomics: Off-targets in epigenome editing

    更新日期:2018-04-04
  • Stem cells: From somatic cells to naive pluripotent cells
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03

    Stem cells: From somatic cells to naive pluripotent cells Stem cells: From somatic cells to naive pluripotent cells, Published online: 03 April 2018; doi:10.1038/nmeth.4648 Stem cells: From somatic cells to naive pluripotent cells

    更新日期:2018-04-04
  • Genomics: Genome editing with ease
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03

    Genomics: Genome editing with ease Genomics: Genome editing with ease, Published online: 03 April 2018; doi:10.1038/nmeth.4652 Genomics: Genome editing with ease

    更新日期:2018-04-04
  • Neuroscience: Minimally invasive optogenetics
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03
    Nina Vogt

    Neuroscience: Minimally invasive optogenetics Neuroscience: Minimally invasive optogenetics, Published online: 03 April 2018; doi:10.1038/nmeth.4654 Upconversion nanoparticles can serve as intermediaries to illuminate optogenetic tools in the mouse brain.

    更新日期:2018-04-04
  • Cell biology: Live-streaming the cytoplasm
    Nat. Methods (IF 25.062) Pub Date : 2018-04-03
    Tal Nawy

    Cell biology: Live-streaming the cytoplasm Cell biology: Live-streaming the cytoplasm, Published online: 03 April 2018; doi:10.1038/nmeth.4656 A new approach uses beams of light to direct cytoplasmic flows.

    更新日期:2018-04-04
  • scmap: projection of single-cell RNA-seq data across data sets
    Nat. Methods (IF 25.062) Pub Date : 2018-04-02
    Vladimir Yu Kiselev, Andrew Yiu, Martin Hemberg

    Single-cell RNA-seq (scRNA-seq) allows researchers to define cell types on the basis of unsupervised clustering of the transcriptome. However, differences in experimental methods and computational analyses make it challenging to compare data across experiments. Here we present scmap (http://bioconductor.org/packages/scmap; web version at http://www.sanger.ac.uk/science/tools/scmap), a method for projecting cells from an scRNA-seq data set onto cell types or individual cells from other experiments.

    更新日期:2018-04-03
  • Real-time fluorescence and deformability cytometry
    Nat. Methods (IF 25.062) Pub Date : 2018-04-02
    Philipp Rosendahl, Katarzyna Plak, Angela Jacobi, Martin Kraeter, Nicole Toepfner, Oliver Otto, Christoph Herold, Maria Winzi, Maik Herbig, Yan Ge, Salvatore Girardo, Katrin Wagner, Buzz Baum, Jochen Guck

    The throughput of cell mechanical characterization has recently approached that of conventional flow cytometers. However, this very sensitive, label-free approach still lacks the specificity of molecular markers. Here we developed an approach that combines real-time 1D-imaging fluorescence and deformability cytometry in one instrument (RT-FDC), thus opening many new research avenues. We demonstrated its utility by using subcellular fluorescence localization to identify mitotic cells and test for mechanical changes in those cells in an RNA interference screen.

    更新日期:2018-04-03
  • Specter: linear deconvolution for targeted analysis of data-independent acquisition mass spectrometry proteomics
    Nat. Methods (IF 25.062) Pub Date : 2018-04-02
    Ryan Peckner, Samuel A Myers, Alvaro Sebastian Vaca Jacome, Jarrett D Egertson, Jennifer G Abelin, Michael J MacCoss, Steven A Carr, Jacob D Jaffe

    Mass spectrometry with data-independent acquisition (DIA) is a promising method to improve the comprehensiveness and reproducibility of targeted and discovery proteomics, in theory by systematically measuring all peptide precursors in a biological sample. However, the analytical challenges involved in discriminating between peptides with similar sequences in convoluted spectra have limited its applicability in important cases, such as the detection of single-nucleotide polymorphisms (SNPs) and alternative site localizations in phosphoproteomics data. We report Specter (https://github.com/rpeckner-broad/Specter), an open-source software tool that uses linear algebra to deconvolute DIA mixture spectra directly through comparison to a spectral library, thus circumventing the problems associated with typical fragment-correlation-based approaches. We validate the sensitivity of Specter and its performance relative to that of other methods, and show that Specter is able to successfully analyze cases involving highly similar peptides that are typically challenging for DIA analysis methods.

    更新日期:2018-04-03
  • Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”
    Nat. Methods (IF 25.062) Pub Date : 2018-03-30
    Reynald M Lescarbeau, Bradley Murray, Thomas M Barnes, Nessan Bermingham

    Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo” Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”, Published online: 30 March 2018; doi:10.1038/nmeth.4553 Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”

    更新日期:2018-03-31
  • Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”
    Nat. Methods (IF 25.062) Pub Date : 2018-03-30
    Lauryl M J Nutter, Jason D Heaney, K C Kent Lloyd, Stephen A Murray, John R Seavitt, William C Skarnes, Lydia Teboul, Steve D M Brown, Mark Moore

    Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo” Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”, Published online: 30 March 2018; doi:10.1038/nmeth.4559 Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”

    更新日期:2018-03-31
  • Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”
    Nat. Methods (IF 25.062) Pub Date : 2018-03-30
    Christopher J Wilson, Tim Fennell, Anne Bothmer, Morgan L Maeder, Deepak Reyon, Cecilia Cotta-Ramusino, Cecilia A Fernandez, Eugenio Marco, Luis A Barrera, Hariharan Jayaram, Charles F Albright, Gerald F Cox, George M Church, Vic E Myer

    Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo” Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”, Published online: 30 March 2018; doi:10.1038/nmeth.4552 Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”

    更新日期:2018-03-31
  • Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”
    Nat. Methods (IF 25.062) Pub Date : 2018-03-30
    Sang-Tae Kim, Jeongbin Park, Daesik Kim, Kyoungmi Kim, Sangsu Bae, Matthias Schlesner, Jin-Soo Kim

    Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo” Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”, Published online: 30 March 2018; doi:10.1038/nmeth.4554 Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”

    更新日期:2018-03-31
  • CRISPR off-targets: a reassessment
    Nat. Methods (IF 25.062) Pub Date : 2018-03-30

    CRISPR off-targets: a reassessment CRISPR off-targets: a reassessment, Published online: 30 March 2018; doi:10.1038/nmeth.4664 There was insufficient data to support the claim of unexpected off-target effects due to CRISPR in a paper published in Nature Methods. More work is needed to determine whether such events occur in vivo.

    更新日期:2018-03-31
  • Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”
    Nat. Methods (IF 25.062) Pub Date : 2018-03-30
    Caleb A Lareau, Kendell Clement, Jonathan Y Hsu, Vikram Pattanayak, J Keith Joung, Martin J Aryee, Luca Pinello

    Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo” Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”, Published online: 30 March 2018; doi:10.1038/nmeth.4541 Response to “Unexpected mutations after CRISPR–Cas9 editing in vivo”

    更新日期:2018-03-31
  • NAMD goes quantum: an integrative suite for hybrid simulations
    Nat. Methods (IF 25.062) Pub Date : 2018-03-26
    Marcelo C R Melo, Rafael C Bernardi, Till Rudack, Maximilian Scheurer, Christoph Riplinger, James C Phillips, Julio D C Maia, Gerd B Rocha, João V Ribeiro, John E Stone, Frank Neese, Klaus Schulten, Zaida Luthey-Schulten

    Hybrid methods that combine quantum mechanics (QM) and molecular mechanics (MM) can be applied to studies of reaction mechanisms in locations ranging from active sites of small enzymes to multiple sites in large bioenergetic complexes. By combining the widely used molecular dynamics and visualization programs NAMD and VMD with the quantum chemistry packages ORCA and MOPAC, we created an integrated, comprehensive, customizable, and easy-to-use suite (http://www.ks.uiuc.edu/Research/qmmm). Through the QwikMD interface, setup, execution, visualization, and analysis are streamlined for all levels of expertise.

    更新日期:2018-03-26
  • Burden-driven feedback control of gene expression
    Nat. Methods (IF 25.062) Pub Date : 2018-03-26
    Francesca Ceroni, Alice Boo, Simone Furini, Thomas E Gorochowski, Olivier Borkowski, Yaseen N Ladak, Ali R Awan, Charlie Gilbert, Guy-Bart Stan, Tom Ellis

    Cells use feedback regulation to ensure robust growth despite fluctuating demands for resources and differing environmental conditions. However, the expression of foreign proteins from engineered constructs is an unnatural burden that cells are not adapted for. Here we combined RNA-seq with an in vivo assay to identify the major transcriptional changes that occur in Escherichia coli when inducible synthetic constructs are expressed. We observed that native promoters related to the heat-shock response activated expression rapidly in response to synthetic expression, regardless of the construct. Using these promoters, we built a dCas9-based feedback-regulation system that automatically adjusts the expression of a synthetic construct in response to burden. Cells equipped with this general-use controller maintained their capacity for native gene expression to ensure robust growth and thus outperformed unregulated cells in terms of protein yield in batch production. This engineered feedback is to our knowledge the first example of a universal, burden-based biomolecular control system and is modular, tunable and portable.

    更新日期:2018-03-26
  • Photoactivatable drugs for nicotinic optopharmacology
    Nat. Methods (IF 25.062) Pub Date : 2018-03-26
    Sambashiva Banala, Matthew C Arvin, Nicholas M Bannon, Xiao-Tao Jin, John J Macklin, Yong Wang, Can Peng, Guiqing Zhao, John J Marshall, Kyle R Gee, David L Wokosin, Veronica J Kim, J Michael McIntosh, Anis Contractor, Henry A Lester, Yevgenia Kozorovitskiy, Ryan M Drenan, Luke D Lavis

    Photoactivatable pharmacological agents have revolutionized neuroscience, but the palette of available compounds is limited. We describe a general method for caging tertiary amines by using a stable quaternary ammonium linkage that elicits a red shift in the activation wavelength. We prepared a photoactivatable nicotine (PA-Nic), uncageable via one- or two-photon excitation, that is useful to study nicotinic acetylcholine receptors (nAChRs) in different experimental preparations and spatiotemporal scales.

    更新日期:2018-03-26
  • A toolbox of immunoprecipitation-grade monoclonal antibodies to human transcription factors
    Nat. Methods (IF 25.062) Pub Date : 2018-03-19
    Anand Venkataraman, Kun Yang, Jose Irizarry, Mark Mackiewicz, Paolo Mita, Zheng Kuang, Lin Xue, Devlina Ghosh, Shuang Liu, Pedro Ramos, Shaohui Hu, Diane Bayron Kain, Sarah Keegan, Richard Saul, Simona Colantonio, Hongyan Zhang, Florencia Pauli Behn, Guang Song, Edisa Albino, Lillyann Asencio, Leonardo Ramos, Luvir Lugo, Gloriner Morell, Javier Rivera, Kimberly Ruiz, Ruth Almodovar, Luis Nazario, Keven Murphy, Ivan Vargas, Zully Ann Rivera-Pacheco, Christian Rosa, Moises Vargas, Jessica McDade, Brian S Clark, Sooyeon Yoo, Seva G Khambadkone, Jimmy de Melo, Milanka Stevanovic, Lizhi Jiang, Yana Li, Wendy Y Yap, Brittany Jones, Atul Tandon, Elliot Campbell, Gaetano T Montelione, Stephen Anderson, Richard M Myers, Jef D Boeke, David Fenyö, Gordon Whiteley, Joel S Bader, Ignacio Pino, Daniel J Eichinger, Heng Zhu, Seth Blackshaw

    A key component of efforts to address the reproducibility crisis in biomedical research is the development of rigorously validated and renewable protein-affinity reagents. As part of the US National Institutes of Health (NIH) Protein Capture Reagents Program (PCRP), we have generated a collection of 1,406 highly validated immunoprecipitation- and/or immunoblotting-grade mouse monoclonal antibodies (mAbs) to 737 human transcription factors, using an integrated production and validation pipeline. We used HuProt human protein microarrays as a primary validation tool to identify mAbs with high specificity for their cognate targets. We further validated PCRP mAbs by means of multiple experimental applications, including immunoprecipitation, immunoblotting, chromatin immunoprecipitation followed by sequencing (ChIP-seq), and immunohistochemistry. We also conducted a meta-analysis that identified critical variables that contribute to the generation of high-quality mAbs. All validation data, protocols, and links to PCRP mAb suppliers are available at http://proteincapture.org.

    更新日期:2018-03-20
  • Identification of spatial expression trends in single-cell gene expression data
    Nat. Methods (IF 25.062) Pub Date : 2018-03-19
    Daniel Edsgärd, Per Johnsson, Rickard Sandberg

    As methods for measuring spatial gene expression at single-cell resolution become available, there is a need for computational analysis strategies. We present trendsceek, a method based on marked point processes that identifies genes with statistically significant spatial expression trends. trendsceek finds these genes in spatial transcriptomic and sequential fluorescence in situ hybridization data, and also reveals significant gene expression gradients and hot spots in low-dimensional projections of dissociated single-cell RNA-seq data.

    更新日期:2018-03-20
  • SpatialDE: identification of spatially variable genes
    Nat. Methods (IF 25.062) Pub Date : 2018-03-19
    Valentine Svensson, Sarah A Teichmann, Oliver Stegle

    Technological advances have made it possible to measure spatially resolved gene expression at high throughput. However, methods to analyze these data are not established. Here we describe SpatialDE, a statistical test to identify genes with spatial patterns of expression variation from multiplexed imaging or spatial RNA-sequencing data. SpatialDE also implements 'automatic expression histology', a spatial gene-clustering approach that enables expression-based tissue histology.

    更新日期:2018-03-20
  • Metagenomic mining of regulatory elements enables programmable species-selective gene expression
    Nat. Methods (IF 25.062) Pub Date : 2018-03-19
    Nathan I Johns, Antonio L C Gomes, Sung Sun Yim, Anthony Yang, Tomasz Blazejewski, Christopher S Smillie, Mark B Smith, Eric J Alm, Sriram Kosuri, Harris H Wang

    Robust and predictably performing synthetic circuits rely on the use of well-characterized regulatory parts across different genetic backgrounds and environmental contexts. Here we report the large-scale metagenomic mining of thousands of natural 5′ regulatory sequences from diverse bacteria, and their multiplexed gene expression characterization in industrially relevant microbes. We identified sequences with broad and host-specific expression properties that are robust in various growth conditions. We also observed substantial differences between species in terms of their capacity to utilize exogenous regulatory sequences. Finally, we demonstrate programmable species-selective gene expression that produces distinct and diverse output patterns in different microbes. Together, these findings provide a rich resource of characterized natural regulatory sequences and a framework that can be used to engineer synthetic gene circuits with unique and tunable cross-species functionality and properties, and also suggest the prospect of ultimately engineering complex behaviors at the community level.

    更新日期:2018-03-20
  • Alignment of single-cell trajectories to compare cellular expression dynamics
    Nat. Methods (IF 25.062) Pub Date : 2018-03-12
    Ayelet Alpert, Lindsay S Moore, Tania Dubovik, Shai S Shen-Orr

    Single-cell RNA sequencing and high-dimensional cytometry can be used to generate detailed trajectories of dynamic biological processes such as differentiation or development. Here we present cellAlign, a quantitative framework for comparing expression dynamics within and between single-cell trajectories. By applying cellAlign to mouse and human embryonic developmental trajectories, we systematically delineate differences in the temporal regulation of gene expression programs that would otherwise be masked.

    更新日期:2018-03-13
  • Improved Ribo-seq enables identification of cryptic translation events
    Nat. Methods (IF 25.062) Pub Date : 2018-03-12
    Florian Erhard, Anne Halenius, Cosima Zimmermann, Anne L'Hernault, Daniel J Kowalewski, Michael P Weekes, Stefan Stevanovic, Ralf Zimmer, Lars Dölken

    Ribosome profiling has been used to predict thousands of short open reading frames (sORFs) in eukaryotic cells, but it suffers from substantial levels of noise. PRICE (https://github.com/erhard-lab/price) is a computational method that models experimental noise to enable researchers to accurately resolve overlapping sORFs and noncanonical translation initiation. We experimentally validated translation using major histocompatibility complex class I (MHC I) peptidomics and observed that sORF-derived peptides efficiently enter the MHC I presentation pathway and thus constitute a substantial fraction of the antigen repertoire.

    更新日期:2018-03-13
  • Using deep learning to model the hierarchical structure and function of a cell
    Nat. Methods (IF 25.062) Pub Date : 2018-03-05
    Jianzhu Ma, Michael Ku Yu, Samson Fong, Keiichiro Ono, Eric Sage, Barry Demchak, Roded Sharan, Trey Ideker

    Although artificial neural networks are powerful classifiers, their internal structures are hard to interpret. In the life sciences, extensive knowledge of cell biology provides an opportunity to design visible neural networks (VNNs) that couple the model's inner workings to those of real systems. Here we develop DCell, a VNN embedded in the hierarchical structure of 2,526 subsystems comprising a eukaryotic cell (http://d-cell.ucsd.edu/). Trained on several million genotypes, DCell simulates cellular growth nearly as accurately as laboratory observations. During simulation, genotypes induce patterns of subsystem activities, enabling in silico investigations of the molecular mechanisms underlying genotype–phenotype associations. These mechanisms can be validated, and many are unexpected; some are governed by Boolean logic. Cumulatively, 80% of the importance for growth prediction is captured by 484 subsystems (21%), reflecting the emergence of a complex phenotype. DCell provides a foundation for decoding the genetics of disease, drug resistance and synthetic life.

    更新日期:2018-03-06
  • Neuroscience: A four-armed patch-clamping robot
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28

    Neuroscience: A four-armed patch-clamping robot Neuroscience: A four-armed patch-clamping robot, Published online: 28 February 2018; doi:10.1038/nmeth.4618 Neuroscience: A four-armed patch-clamping robot

    更新日期:2018-03-01
  • Systems biology: Tracking the protein–metabolite interactome
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28
    Allison Doerr

    Systems biology: Tracking the protein–metabolite interactome Systems biology: Tracking the protein–metabolite interactome, Published online: 28 February 2018; doi:10.1038/nmeth.4622 A method combining limited proteolysis with mass spectrometry systematically detects protein–metabolite interactions.

    更新日期:2018-03-01
  • Genomics: Imaging allelic loci in live cells
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28
    Nina Vogt

    Genomics: Imaging allelic loci in live cells Genomics: Imaging allelic loci in live cells, Published online: 28 February 2018; doi:10.1038/nmeth.4624 SNP-CLING enables determining the dynamics of alleles on different chromosomes in live cells.

    更新日期:2018-03-01
  • The Author File: Miguel Angel Esteban
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28
    Vivien Marx

    The Author File: Miguel Angel Esteban The Author File: Miguel Angel Esteban, Published online: 28 February 2018; doi:10.1038/nmeth.4626 A way to study the RNA interactome, baby RNA encounters, a sprinkle of poetry.

    更新日期:2018-03-01
  • Sharing epigenomes globally
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28

    Sharing epigenomes globally Sharing epigenomes globally, Published online: 28 February 2018; doi:10.1038/nmeth.4630 Epigenome reference data are continually being enriched—researchers should explore them, even if raw data access still presents some hurdles.

    更新日期:2018-03-01
  • Picky: a simple online PRM and SRM method designer for targeted proteomics
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28
    Henrik Zauber, Marieluise Kirchner, Matthias Selbach

    Picky: a simple online PRM and SRM method designer for targeted proteomics Picky: a simple online PRM and SRM method designer for targeted proteomics, Published online: 28 February 2018; doi:10.1038/nmeth.4607 Picky: a simple online PRM and SRM method designer for targeted proteomics

    更新日期:2018-03-01
  • Erratum: Depleting endogenous proteins
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28
    Ada Yee

    Erratum: Depleting endogenous proteins Erratum: Depleting endogenous proteins, Published online: 28 February 2018; doi:10.1038/nmeth0318-226a Erratum: Depleting endogenous proteins

    更新日期:2018-03-01
  • Putting microfluidics in other people's hands
    Nat. Methods (IF 25.062) Pub Date : 2018-02-28
    Vivien Marx

    Putting microfluidics in other people's hands Putting microfluidics in other people's hands, Published online: 28 February 2018; doi:10.1038/nmeth.4609 In microfluidics, sharing is hard. But practitioners are exploring new ways to share designs, devices and experience.

    更新日期:2018-03-01
  • A hybridization-chain-reaction-based method for amplifying immunosignals
    Nat. Methods (IF 25.062) Pub Date : 2018-02-26
    Rui Lin, Qiru Feng, Peng Li, Ping Zhou, Ruiyu Wang, Zhe Liu, Zhiqiang Wang, Xiangbing Qi, Nan Tang, Feng Shao, Minmin Luo

    Immunosignal hybridization chain reaction (isHCR) combines antibody–antigen interactions with hybridization chain reaction (HCR) technology, which results in amplification of immunofluorescence signals by up to two to three orders of magnitude with low background. isHCR's highly modular and easily adaptable design enables the technique to be applied broadly, and we further optimized its use in multiplexed imaging and in state-of-the-art tissue expansion and clearing techniques.

    更新日期:2018-02-27
  • Cell-type specific sequencing of microRNAs from complex animal tissues
    Nat. Methods (IF 25.062) Pub Date : 2018-02-26
    Chiara Alberti, Raphael A Manzenreither, Ivica Sowemimo, Thomas R Burkard, Jingkui Wang, Katharina Mahofsky, Stefan L Ameres, Luisa Cochella

    MicroRNAs (miRNAs) play an essential role in the post-transcriptional regulation of animal development and physiology. However, in vivo studies aimed at linking miRNA function to the biology of distinct cell types within complex tissues remain challenging, partly because in vivo miRNA-profiling methods lack cellular resolution. We report microRNome by methylation-dependent sequencing (mime-seq), an in vivo enzymatic small-RNA-tagging approach that enables high-throughput sequencing of tissue- and cell-type-specific miRNAs in animals. The method combines cell-type-specific 3′-terminal 2′-O-methylation of animal miRNAs by a genetically encoded, plant-specific methyltransferase (HEN1), with chemoselective small-RNA cloning and high-throughput sequencing. We show that mime-seq uncovers the miRNomes of specific cells within Caenorhabditis elegans and Drosophila at unprecedented specificity and sensitivity, enabling miRNA profiling with single-cell resolution in whole animals. Mime-seq overcomes current challenges in cell-type-specific small-RNA profiling and provides novel entry points for understanding the function of miRNAs in spatially restricted physiological settings.

    更新日期:2018-02-27
  • Bias, robustness and scalability in single-cell differential expression analysis
    Nat. Methods (IF 25.062) Pub Date : 2018-02-26
    Charlotte Soneson, Mark D Robinson

    Many methods have been used to determine differential gene expression from single-cell RNA (scRNA)-seq data. We evaluated 36 approaches using experimental and synthetic data and found considerable differences in the number and characteristics of the genes that are called differentially expressed. Prefiltering of lowly expressed genes has important effects, particularly for some of the methods developed for bulk RNA-seq data analysis. However, we found that bulk RNA-seq analysis methods do not generally perform worse than those developed specifically for scRNA-seq. We also present conquer, a repository of consistently processed, analysis-ready public scRNA-seq data sets that is aimed at simplifying method evaluation and reanalysis of published results. Each data set provides abundance estimates for both genes and transcripts, as well as quality control and exploratory analysis reports.

    更新日期:2018-02-27
  • Quantitative mapping and minimization of super-resolution optical imaging artifacts
    Nat. Methods (IF 25.062) Pub Date : 2018-02-19
    Siân Culley, David Albrecht, Caron Jacobs, Pedro Matos Pereira, Christophe Leterrier, Jason Mercer, Ricardo Henriques

    Super-resolution microscopy depends on steps that can contribute to the formation of image artifacts, leading to misinterpretation of biological information. We present NanoJ-SQUIRREL, an ImageJ-based analytical approach that provides quantitative assessment of super-resolution image quality. By comparing diffraction-limited images and super-resolution equivalents of the same acquisition volume, this approach generates a quantitative map of super-resolution defects and can guide researchers in optimizing imaging parameters.

    更新日期:2018-02-21
  • On the design of CRISPR-based single-cell molecular screens
    Nat. Methods (IF 25.062) Pub Date : 2018-02-19
    Andrew J Hill, José L McFaline-Figueroa, Lea M Starita, Molly J Gasperini, Kenneth A Matreyek, Jonathan Packer, Dana Jackson, Jay Shendure, Cole Trapnell

    Several groups recently coupled CRISPR perturbations and single-cell RNA-seq for pooled genetic screens. We demonstrate that vector designs of these studies are susceptible to ∼50% swapping of guide RNA–barcode associations because of lentiviral template switching. We optimized a published alternative, CROP-seq, in which the guide RNA also serves as the barcode, and here confirm that this strategy performs robustly and doubled the rate at which guides are assigned to cells to 94%.

    更新日期:2018-02-21
  • Identifying the favored mutation in a positive selective sweep
    Nat. Methods (IF 25.062) Pub Date : 2018-02-19
    Ali Akbari, Joseph J Vitti, Arya Iranmehr, Mehrdad Bakhtiari, Pardis C Sabeti, Siavash Mirarab, Vineet Bafna

    Most approaches that capture signatures of selective sweeps in population genomics data do not identify the specific mutation favored by selection. We present iSAFE (for “integrated selection of allele favored by evolution”), a method that enables researchers to accurately pinpoint the favored mutation in a large region (∼5 Mbp) by using a statistic derived solely from population genetics signals. iSAFE does not require knowledge of demography, the phenotype under selection, or functional annotations of mutations.

    更新日期:2018-02-21
  • Capturing the interactome of newly transcribed RNA
    Nat. Methods (IF 25.062) Pub Date : 2018-02-12
    Xichen Bao, Xiangpeng Guo, Menghui Yin, Muqddas Tariq, Yiwei Lai, Shahzina Kanwal, Jiajian Zhou, Na Li, Yuan Lv, Carlos Pulido-Quetglas, Xiwei Wang, Lu Ji, Muhammad J Khan, Xihua Zhu, Zhiwei Luo, Changwei Shao, Do-Hwan Lim, Xiao Liu, Nan Li, Wei Wang, Minghui He, Yu-Lin Liu, Carl Ward, Tong Wang, Gong Zhang, Dongye Wang, Jianhua Yang, Yiwen Chen, Chaolin Zhang, Ralf Jauch, Yun-Gui Yang, Yangming Wang, Baoming Qin, Minna-Liisa Anko, Andrew P Hutchins, Hao Sun, Huating Wang, Xiang-Dong Fu, Biliang Zhang, Miguel A Esteban

    We combine the labeling of newly transcribed RNAs with 5-ethynyluridine with the characterization of bound proteins. This approach, named capture of the newly transcribed RNA interactome using click chemistry (RICK), systematically captures proteins bound to a wide range of RNAs, including nascent RNAs and traditionally neglected nonpolyadenylated RNAs. RICK has identified mitotic regulators amongst other novel RNA-binding proteins with preferential affinity for nonpolyadenylated RNAs, revealed a link between metabolic enzymes/factors and nascent RNAs, and expanded the known RNA-bound proteome of mouse embryonic stem cells. RICK will facilitate an in-depth interrogation of the total RNA-bound proteome in different cells and systems.

    更新日期:2018-02-13
  • RNA–protein interaction detection in living cells
    Nat. Methods (IF 25.062) Pub Date : 2018-02-05
    Muthukumar Ramanathan, Karim Majzoub, Deepti S Rao, Poornima H Neela, Brian J Zarnegar, Smarajit Mondal, Julien G Roth, Hui Gai, Joanna R Kovalski, Zurab Siprashvili, Theo D Palmer, Jan E Carette, Paul A Khavari

    RNA–protein interactions play numerous roles in cellular function and disease. Here we describe RNA–protein interaction detection (RaPID), which uses proximity-dependent protein labeling, based on the BirA* biotin ligase, to rapidly identify the proteins that bind RNA sequences of interest in living cells. RaPID displays utility in multiple applications, including in evaluating protein binding to mutant RNA motifs in human genetic disorders, in uncovering potential post-transcriptional networks in breast cancer, and in discovering essential host proteins that interact with Zika virus RNA. To improve the BirA*-labeling component of RaPID, moreover, a new mutant BirA* was engineered from Bacillus subtilis, termed BASU, that enables >1,000-fold faster kinetics and >30-fold increased signal-to-noise ratio over the prior standard Escherichia coli BirA*, thereby enabling direct study of RNA–protein interactions in living cells on a timescale as short as 1 min.

    更新日期:2018-02-06
  • The Author File: Jennifer Phillips-Cremins
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30
    Vivien Marx

    The Author File: Jennifer Phillips-Cremins The Author File: Jennifer Phillips-Cremins, Published online: 30 January 2018; doi:10.1038/nmeth.4584 To better explore how genomes fold takes disparate fields and a love of math.

    更新日期:2018-01-30
  • Neuroscience: Myelin quantification at nanoscale
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30

    Neuroscience: Myelin quantification at nanoscale Neuroscience: Myelin quantification at nanoscale, Published online: 30 January 2018; doi:10.1038/nmeth.4586 Neuroscience: Myelin quantification at nanoscale

    更新日期:2018-01-30
  • Imaging: A colorful series of bioorthogonal probes
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30

    Imaging: A colorful series of bioorthogonal probes Imaging: A colorful series of bioorthogonal probes, Published online: 30 January 2018; doi:10.1038/nmeth.4590 Imaging: A colorful series of bioorthogonal probes

    更新日期:2018-01-30
  • Imaging: Single-molecule imaging and force spectroscopy at extended depth
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30

    Imaging: Single-molecule imaging and force spectroscopy at extended depth Imaging: Single-molecule imaging and force spectroscopy at extended depth, Published online: 30 January 2018; doi:10.1038/nmeth.4588 Imaging: Single-molecule imaging and force spectroscopy at extended depth

    更新日期:2018-01-30
  • Proteomics: Cell-type-specific proteomics in the in vivo mouse brain
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30

    Proteomics: Cell-type-specific proteomics in the in vivo mouse brain Proteomics: Cell-type-specific proteomics in the in vivo mouse brain, Published online: 30 January 2018; doi:10.1038/nmeth.4592 Proteomics: Cell-type-specific proteomics in the in vivo mouse brain

    更新日期:2018-01-30
  • Meet some code-breakers of noncoding RNAs
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30
    Vivien Marx

    Meet some code-breakers of noncoding RNAs Meet some code-breakers of noncoding RNAs, Published online: 30 January 2018; doi:10.1038/nmeth.4594 The regulome—the part of the genome that regulates function—includes noncoding RNAs with varied functions yet to be deciphered.

    更新日期:2018-01-30
  • Neuroscience: Putting a stamp on single cells
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30
    Nina Vogt

    Neuroscience: Putting a stamp on single cells Neuroscience: Putting a stamp on single cells, Published online: 30 January 2018; doi:10.1038/nmeth.4596 Virus stamping can target single cells in complex tissues, both in culture and in vivo.

    更新日期:2018-01-30
  • Epigenetics: Humanized yeast—erasing 1.3 billion years of evolution
    Nat. Methods (IF 25.062) Pub Date : 2018-01-30
    Nicole Rusk

    Epigenetics: Humanized yeast—erasing 1.3 billion years of evolution Epigenetics: Humanized yeast—erasing 1.3 billion years of evolution, Published online: 30 January 2018; doi:10.1038/nmeth.4598 Cellular engineering that allows budding yeast to survive with the four core human histones opens the door to exploring the function of histone variants and their modifications.

    更新日期:2018-01-30
  • Ab initio electron density determination directly from solution scattering data
    Nat. Methods (IF 25.062) Pub Date : 2018-01-29
    Thomas D Grant

    Using a novel iterative structure factor retrieval algorithm, here I show that electron density can be directly calculated from solution scattering data without modeling. The algorithm was validated with experimental data from 12 different biological macromolecules. This approach avoids many of the assumptions limiting the resolution and accuracy of modeling algorithms by explicitly calculating electron density. This algorithm can be applied to a wide variety of molecular systems.

    更新日期:2018-01-29
  • STED super-resolved microscopy
    Nat. Methods (IF 25.062) Pub Date : 2018-01-29
    Giuseppe Vicidomini, Paolo Bianchini, Alberto Diaspro

    Stimulated emission depletion (STED) microscopy provides subdiffraction resolution while preserving useful aspects of fluorescence microscopy, such as optical sectioning, and molecular specificity and sensitivity. However, sophisticated microscopy architectures and high illumination intensities have limited STED microscopy's widespread use in the past. Here we summarize the progress that is mitigating these problems and giving substantial momentum to STED microscopy applications. We discuss the future of this method in regard to spatiotemporal limits, live-cell imaging and combination with spectroscopy. Advances in these areas may elevate STED microscopy to a standard method for imaging in the life sciences.

    更新日期:2018-01-29
  • Widespread bacterial protein histidine phosphorylation revealed by mass spectrometry-based proteomics
    Nat. Methods (IF 25.062) Pub Date : 2018-01-29
    Clement M Potel, Miao-Hsia Lin, Albert J R Heck, Simone Lemeer

    For decades, major difficulties in analyzing histidine phosphorylation have limited the study of phosphohistidine signaling. Here we report a method revealing widespread and abundant protein histidine phosphorylation in Escherichia coli. We generated an extensive E. coli phosphoproteome data set, in which a remarkably high percentage (∼10%) of phosphorylation sites are phosphohistidine sites. This resource should help enable a better understanding of the biological function of histidine phosphorylation.

    更新日期:2018-01-29
  • TimeLapse-seq: adding a temporal dimension to RNA sequencing through nucleoside recoding
    Nat. Methods (IF 25.062) Pub Date : 2018-01-22
    Jeremy A Schofield, Erin E Duffy, Lea Kiefer, Meaghan C Sullivan, Matthew D Simon

    RNA sequencing (RNA-seq) offers a snapshot of cellular RNA populations, but not temporal information about the sequenced RNA. Here we report TimeLapse-seq, which uses oxidative-nucleophilic-aromatic substitution to convert 4-thiouridine into cytidine analogs, yielding apparent U-to-C mutations that mark new transcripts upon sequencing. TimeLapse-seq is a single-molecule approach that is adaptable to many applications and reveals RNA dynamics and induced differential expression concealed in traditional RNA-seq.

    更新日期:2018-01-23
  • EVIR: chimeric receptors that enhance dendritic cell cross-dressing with tumor antigens
    Nat. Methods (IF 25.062) Pub Date : 2018-01-22
    Mario Leonardo Squadrito, Chiara Cianciaruso, Sarah K Hansen, Michele De Palma

    We describe a lentivirus-encoded chimeric receptor, termed extracellular vesicle (EV)-internalizing receptor (EVIR), which enables the selective uptake of cancer-cell-derived EVs by dendritic cells (DCs). The EVIR enhances DC presentation of EV-associated tumor antigens to CD8+ T cells primarily through MHCI recycling and cross-dressing. EVIRs should facilitate exploring the mechanisms and implications of horizontal transfer of tumor antigens to antigen-presenting cells.

    更新日期:2018-01-22
  • Highly parallel direct RNA sequencing on an array of nanopores
    Nat. Methods (IF 25.062) Pub Date : 2018-01-15
    Daniel R Garalde, Elizabeth A Snell, Daniel Jachimowicz, Botond Sipos, Joseph H Lloyd, Mark Bruce, Nadia Pantic, Tigist Admassu, Phillip James, Anthony Warland, Michael Jordan, Jonah Ciccone, Sabrina Serra, Jemma Keenan, Samuel Martin, Luke McNeill, E Jayne Wallace, Lakmal Jayasinghe, Chris Wright, Javier Blasco, Stephen Young, Denise Brocklebank, Sissel Juul, James Clarke, Andrew J Heron, Daniel J Turner

    Sequencing the RNA in a biological sample can unlock a wealth of information, including the identity of bacteria and viruses, the nuances of alternative splicing or the transcriptional state of organisms. However, current methods have limitations due to short read lengths and reverse transcription or amplification biases. Here we demonstrate nanopore direct RNA-seq, a highly parallel, real-time, single-molecule method that circumvents reverse transcription or amplification steps. This method yields full-length, strand-specific RNA sequences and enables the direct detection of nucleotide analogs in RNA.

    更新日期:2018-01-15
  • Detecting hierarchical genome folding with network modularity
    Nat. Methods (IF 25.062) Pub Date : 2018-01-15
    Heidi K Norton, Daniel J Emerson, Harvey Huang, Jesi Kim, Katelyn R Titus, Shi Gu, Danielle S Bassett, Jennifer E Phillips-Cremins

    Mammalian genomes are folded in a hierarchy of compartments, topologically associating domains (TADs), subTADs and looping interactions. Here, we describe 3DNetMod, a graph theory-based method for sensitive and accurate detection of chromatin domains across length scales in Hi-C data. We identify nested, partially overlapping TADs and subTADs genome wide by optimizing network modularity and varying a single resolution parameter. 3DNetMod can be applied broadly to understand genome reconfiguration in development and disease.

    更新日期:2018-01-15
Some contents have been Reproduced with permission of the American Chemical Society.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
化学 • 材料 期刊列表