Table of contents


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Editorial

Kick the bar chart habit p113

doi:10.1038/nmeth.2837

Bar charts are too frequently used to communicate data that they cannot represent well. We strongly encourage the use of more appropriate plots to display statistical samples.


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This Month

The Author File: Janet Thornton p115

Vivien Marx

doi:10.1038/nmeth.2831

Finding ways to navigate the reactions of life and herd tigers are all part of her workday.


Points of View: Bar charts and box plots p117

Marc Streit & Nils Gehlenborg

doi:10.1038/nmeth.2807

Creating a simple yet effective plot requires an understanding of data and tasks.


Points of Significance: Visualizing samples with box plots pp119 - 120

Martin Krzywinski & Naomi Altman

doi:10.1038/nmeth.2813

Use box plots to illustrate the spread and differences of samples.


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Correspondence

BoxPlotR: a web tool for generation of box plots pp121 - 122

Michaela Spitzer, Jan Wildenhain, Juri Rappsilber & Mike Tyers

doi:10.1038/nmeth.2811


E-CRISP: fast CRISPR target site identification pp122 - 123

Florian Heigwer, Grainne Kerr & Michael Boutros

doi:10.1038/nmeth.2812


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Research Highlights

Screening CRISPly in human cells p125

Two groups demonstrate the use of the CRISPR-Cas9 system for genetic screens in human cells.

Zooming in on nuclear logistics pp126 - 127

Cryo-electron tomography, single-particle analysis and cross-linking mass spectroscopy join forces to solve the nuclear pore scaffold puzzle.

Locating the kiss of death pp126 - 127

Chromatin immunoprecipitation of transcription factors tagged for degradation reveals how protein turnover regulates transcription.

A genetic handle on brain circuits p128

Cre-recombinase mouse driver lines provide tools for the functional dissection of neuronal circuits.

Orthogonal logic gates p132

A set of 16 TetR proteins specific for their synthetic promoters will allow the construction of complex genetic circuits.

Methods in Brief

Tools in Brief

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Technology Feature

A blooming genomic desert pp135 - 138

Vivien Marx

doi:10.1038/nmeth.2817

Identifying functional regions in genomes takes scientists beyond protein-coding regions and into stretches formerly known as genomic deserts.


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News and Views

Twitching towards the ideal calcium sensor pp139 - 140

Christian D Wilms & Michael Häusser

doi:10.1038/nmeth.2814

A new family of genetically encoded ratiometric calcium indicators optimized for imaging of calcium signals in vivo exhibits near-linear fluorescence dynamics while minimizing artifacts caused by movement.

See also: Article by Thestrup et al.


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Perspective

The potential of optofluidic biolasers pp141 - 147

Xudong Fan & Seok-Hyun Yun

doi:10.1038/nmeth.2805

Optofluidic biolasers are emerging as a highly sensitive way to measure changes in biological molecules. Biolasers, which incorporate biological material into the gain medium and contain an optical cavity in a fluidic environment, take advantage of the amplification that occurs during laser generation to quantify tiny changes in biological processes in the gain medium. This Perspective describes the principle of the optofluidic biolaser, reviews recent progress and provides outlooks on potential applications and directions for developing this enabling technology.


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Analysis

Demonstrating the feasibility of large-scale development of standardized assays to quantify human proteins pp149 - 155

Jacob J Kennedy, Susan E Abbatiello, Kyunggon Kim, Ping Yan, Jeffrey R Whiteaker, Chenwei Lin, Jun Seok Kim, Yuzheng Zhang, Xianlong Wang, Richard G Ivey, Lei Zhao, Hophil Min, Youngju Lee, Myeong-Hee Yu, Eun Gyeong Yang, Cheolju Lee, Pei Wang, Henry Rodriguez, Youngsoo Kim, Steven A Carr & Amanda G Paulovich

doi:10.1038/nmeth.2763

This Analysis reports the development and assessment of 645 multiple reaction monitoring (MRM) mass spectrometry assays to quantify 319 targeted human breast cancer proteins. The results of this pilot project coordinated among three individual groups suggest that an organized international effort to generate MRM assays to the human proteome will be possible.


Single-molecule evaluation of fluorescent protein photoactivation efficiency using an in vivo nanotemplate pp156 - 162

Nela Durisic, Lara Laparra-Cuervo, Ángel Sandoval-Álvarez, Joseph Steven Borbely & Melike Lakadamyali

doi:10.1038/nmeth.2784

A system using the human glycine receptor expressed in Xenopus oocytes allows characterization of the photoactivation efficiency of photoactivatable and photoconvertible fluorescent proteins at the single-molecule level, providing crucial data for using these probes for quantitative super-resolution microscopy.


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Brief Communications

Quantitative single-cell RNA-seq with unique molecular identifiers pp163 - 166

Saiful Islam, Amit Zeisel, Simon Joost, Gioele La Manno, Pawel Zajac, Maria Kasper, Peter Lönnerberg & Sten Linnarsson

doi:10.1038/nmeth.2772

With an optimized protocol and unique molecular identifiers (UMIs) to tag individual transcripts, the mRNA complement of a single cell can be quantified on an absolute scale with almost no amplification bias.


Drift time-specific collision energies enable deep-coverage data-independent acquisition proteomics pp167 - 170

Ute Distler, Jörg Kuharev, Pedro Navarro, Yishai Levin, Hansjörg Schild & Stefan Tenzer

doi:10.1038/nmeth.2767

A data-independent acquisition (DIA) mass spectrometry approach, ultradefinition (UD)MSE, offers high reproducibility and improved proteome coverage over alternative DIA and data-dependent acquisition workflows.


EC-BLAST: a tool to automatically search and compare enzyme reactions pp171 - 174

Syed Asad Rahman, Sergio Martinez Cuesta, Nicholas Furnham, Gemma L Holliday & Janet M Thornton

doi:10.1038/nmeth.2803

EC-BLAST is a Web-based tool allowing quantitative similarity searches between enzymes at the levels of bond-change, reaction-center or reaction-structure similarity. The tool may help improve the annotation of enzyme function.


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Articles

Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T lymphocytes pp175 - 182

Thomas Thestrup, Julia Litzlbauer, Ingo Bartholomäus, Marsilius Mues, Luigi Russo, Hod Dana, Yuri Kovalchuk, Yajie Liang, Georgios Kalamakis, Yvonne Laukat, Stefan Becker, Gregor Witte, Anselm Geiger, Taylor Allen, Lawrence C Rome, Tsai-Wen Chen, Douglas S Kim, Olga Garaschuk, Christian Griesinger & Oliver Griesbeck

doi:10.1038/nmeth.2773

A collection of improved FRET-based calcium biosensors, called Twitch sensors, is described. Twitches have a reduced number of calcium binding sites per sensor and display high sensitivity in in vivo imaging experiments in mouse brain and lymph node.

See also: News and Views by Wilms & Häusser


Quantifying cell-generated mechanical forces within living embryonic tissues pp183 - 189

Otger Campàs, Tadanori Mammoto, Sean Hasso, Ralph A Sperling, Daniel O'Connell, Ashley G Bischof, Richard Maas, David A Weitz, L Mahadevan & Donald E Ingber

doi:10.1038/nmeth.2761

Microscopic oil droplets with defined mechanical properties are used to measure forces exerted within living tissue.


Transcriptome in vivo analysis (TIVA) of spatially defined single cells in live tissue pp190 - 196

Ditte Lovatt, Brittani K Ruble, Jaehee Lee, Hannah Dueck, Tae Kyung Kim, Stephen Fisher, Chantal Francis, Jennifer M Spaethling, John A Wolf, M Sean Grady, Alexandra V Ulyanova, Sean B Yeldell, Julianne C Griepenburg, Peter T Buckley, Junhyong Kim, Jai-Yoon Sul, Ivan J Dmochowski & James Eberwine

doi:10.1038/nmeth.2804

A noninvasive method is reported for the isolation and profiling of the transcriptome from a single cell in the context of intact tissue.


Scalable inference of heterogeneous reaction kinetics from pooled single-cell recordings pp197 - 202

Christoph Zechner, Michael Unger, Serge Pelet, Matthias Peter & Heinz Koeppl

doi:10.1038/nmeth.2794

This paper describes a method for statistical inference of unobserved molecular states from single-cell time-lapse data.


High-resolution mapping of transcription factor binding sites on native chromatin pp203 - 209

Sivakanthan Kasinathan, Guillermo A Orsi, Gabriel E Zentner, Kami Ahmad & Steven Henikoff

doi:10.1038/nmeth.2766

A method to map transcription factor binding across the genome, at high resolution and without cross-linking.


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Corrigenda

Corrigendum: Functional labeling of neurons and their projections using the synthetic activity–dependent promoter E-SARE p210

Takashi Kawashima, Kazuo Kitamura, Kanzo Suzuki, Mio Nonaka, Satoshi Kamijo, Sayaka Takemoto-Kimura, Masanobu Kano, Hiroyuki Okuno, Kenichi Ohki & Haruhiko Bito

doi:10.1038/nmeth0214-210a


Corrigendum: Accounting for technical noise in single-cell RNA-seq experiments p210

Philip Brennecke, Simon Anders, Jong Kyoung Kim, Aleksandra A Kołodziejczyk, Xiuwei Zhang, Valentina Proserpio, Bianka Baying, Vladimir Benes, Sarah A Teichmann, John C Marioni & Marcus G Heisler

doi:10.1038/nmeth0214-210b


Corrigendum: Comparative analysis of RNA sequencing methods for degraded or low-input samples p210

Xian Adiconis, Diego Borges-Rivera, Rahul Satija, David S DeLuca, Michele A Busby, Aaron M Berlin, Andrey Sivachenko, Dawn Anne Thompson, Alec Wysoker, Timothy Fennell, Andreas Gnirke, Nathalie Pochet, Aviv Regev & Joshua Z Levin

doi:10.1038/nmeth0214-210e


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Errata

Erratum: Adding an unnatural covalent bond to proteins through proximity-enhanced bioreactivity p210

Zheng Xiang, Haiyan Ren, Ying S Hu, Irene Coin, Jing Wei, Hu Cang & Lei Wang

doi:10.1038/nmeth0214-210c


Erratum: Power and sample size p210

Martin Krzywinski & Naomi Altman

doi:10.1038/nmeth0214-210d


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