Table of contents

Method of the Year 2011

Nature Methods' choice for Method of the Year 2011 is genome editing with engineered nucleases. This collection of articles—and the related video—highlights how the ability to use engineered nucleases to make precise, tailored and specific changes to coding and noncoding sequences of the genome, in cells and in organisms of many species, could revolutionize the study of gene function. The Methods to Watch bring together possible future choices for Method of the Year.


Special Feature



Method of the Year 2011 p1


The ability to introduce targeted, tailored changes into the genomes of several species will make it feasible to ask more precise biological questions.


This Month

The author file: Khalid Salaita p3

Monya Baker


Measuring single-molecule forces with light.

Points of view: Data exploration p5

Noam Shoresh & Bang Wong


Enhancement of pattern discovery through graphical representation of data.



GeneProf: analysis of high-throughput sequencing experiments pp7 - 8

Florian Halbritter, Harsh J Vaidya & Simon R Tomlinson


Gene expression deconvolution in linear space pp8 - 9

Yi Zhong & Zhandong Liu


Gene expression deconvolution in linear space p9

Shai S Shen-Orr, Robert Tibshirani & Atul J Butte


Optimal enzymes for amplifying sequencing libraries pp10 - 11

Michael A Quail, Thomas D Otto, Yong Gu, Simon R Harris, Thomas F Skelly, Jacqueline A McQuillan, Harold P Swerdlow & Samuel O Oyola



Research Highlights

Building better bubbles p13

Synthetic clusters of membrane-bound droplets may provide a useful means for simulating tissues or transporting therapeutic payloads.

Three-dimensional genetics pp14 - 15

A combination of chromosome conformation capture carbon copy (5C), modeling and automated imaging renders an empirical three-dimensional model of a bacterial genome.

Peering deeper into the transcriptome pp14 - 15

Targeting portions of the transcriptome for deep sequencing reveals very rare transcripts.

The survival of the fittest p16

An analysis of over 100 human embryonic stem cell lines reveals a genetic change that might confer a growth advantage.

Organic chemistry expands imaging options p19

A new fluorination approach enables the synthesis of diverse PET probes.

A handle on neurodegenerative disease complexity p21

Combining experiments and calculations makes it possible to measure the prognostic value of toxic protein species in the cell.


News Feature

Focus on Method of the Year 2011

Gene-editing nucleases pp23 - 26

Monya Baker


Precise ways to modify the genome arose from unexpected places. Monya Baker reports.



Focus on Method of the Year 2011

Primer: genome editing with engineered nucleases p27

Natalie de Souza


A brief description of tools for targeted cleavage and tailored modification of genomes is presented.



Focus on Method of the Year 2011

Gene editing: not just for translation anymore pp28 - 31

Moira A McMahon, Meghdad Rahdar & Matthew Porteus


Engineered nucleases have advanced the field of gene therapy with the promise of targeted genome modification as a treatment for human diseases. Here we discuss why engineered nucleases are an exciting research tool for gene editing and consider their applications to a range of biological questions.

Focus on Method of the Year 2011

Zinc-finger nucleases: how to play two good hands pp32 - 34

Mark Isalan


Zinc-finger nuclease dimers are more difficult to engineer than single DNA-binding domains, but the development of new methods could help.


Methods to Watch

Focus on Method of the Year 2011

Single-cell methods p35

Natalie de Souza


Improved single-cell methods are helping to unravel biological complexity.

Focus on Method of the Year 2011

Functional genomic resources p35

Nicole Rusk


Tools to manipulate murine genes on a genome-wide scale and to phenotype their effects in animals are maturing.

Focus on Method of the Year 2011

Glycoproteomics p36

Allison Doerr


Methods for tackling the enormously complex glycoproteome are sorely needed.

Focus on Method of the Year 2011

Causal mutations in a haploid landscape p36

Nicole Rusk


Sequencing a haploid genome and understanding the impact of its variants requires technical and computational improvements.

Focus on Method of the Year 2011

Imaging life with thin sheets of light p37

Erika Pastrana


The revival of light-sheet microscopy opens new possibilities for the imaging of living processes.

Focus on Method of the Year 2011

Non–model organisms p37

Tal Nawy


Next-generation sequencing is broadening the application of genetic and genomic studies to the panoply of life.

Focus on Method of the Year 2011

Light-based electrophysiology p38

Erika Pastrana


Genetically encoded voltage sensors are finally measuring up.

Focus on Method of the Year 2011

RNA structures p38

Petya V Krasteva


Accurate methods for RNA-structure determination are being developed.


Technology Feature

Quantitative data: learning to share pp39 - 41

Monya Baker


Adaptive technologies are helping researchers combine and organize experimental results.


News and Views

Running in reverse: rhodopsins sense voltage pp43 - 44

Loren L Looger


Microbial rhodopsins convert light into ion flux; in neurons, this can be used to control activity. New work shows that the opposite is also true: rhodopsins can be used to visualize neural activity.

See also: Article by Kralj et al.

Tracking genomic hydroxymethylation by the base pp45 - 46

Gilles Salbert & Michael Weber


A method uses single-molecule, real-time DNA sequencing to detect the modified base 5-hydroxymethylcytosine, an epigenetic mark recently suspected of having essential roles in genome regulation.

See also: Brief Communication by Song et al.




Power tools for gene expression and clonal analysis in Drosophila pp47 - 55

Alberto del Valle Rodríguez, Dominic Didiano & Claude Desplan


This Review covers recent technological developments to label and manipulate genes in selected populations of cells in Drosophila melanogaster. The Review is intended as a user guide to help with the selection of the best expression systems and clonal analysis techniques for developmental studies in the fly.



A guide to analysis of mouse energy metabolism pp57 - 63

Matthias H Tschöp, John R Speakman, Jonathan R S Arch, Johan Auwerx, Jens C Brüning, Lawrence Chan, Robert H Eckel, Robert V Farese Jr, Jose E Galgani, Catherine Hambly, Mark A Herman, Tamas L Horvath, Barbara B Kahn, Sara C Kozma, Eleftheria Maratos-Flier, Timo D Müller, Heike Münzberg, Paul T Pfluger, Leona Plum, Marc L Reitman, Kamal Rahmouni, Gerald I Shulman, George Thomas, C Ronald Kahn & Eric Ravussin


Authors present workflows for the analysis of metabolism phenotypes in mice and recommend analysis of covariance to asses body composition effects.


Brief Communications

Visualizing mechanical tension across membrane receptors with a fluorescent sensor pp64 - 67

Daniel R Stabley, Carol Jurchenko, Stephen S Marshall & Khalid S Salaita


A fluorescent molecular tension sensor for spatially and temporally mapping the mechanical strain exerted by cell-surface receptors in living cells is described.

Cyanine fluorophore derivatives with enhanced photostability pp68 - 71

Roger B Altman, Daniel S Terry, Zhou Zhou, Qinsi Zheng, Peter Geggier, Rachel A Kolster, Yongfang Zhao, Jonathan A Javitch, J David Warren & Scott C Blanchard


Conjugation of triplet-state quenchers to the small organic cyanine fluorophore, Cy5, increases photostability without affecting its spectral characteristics. This allows longer fluorescence imaging with a concomitant reduction in blinking both in vitro and in living cells.

Counting absolute numbers of molecules using unique molecular identifiers pp72 - 74

Teemu Kivioja, Anna Vähärautio, Kasper Karlsson, Martin Bonke, Martin Enge, Sten Linnarsson & Jussi Taipale


Unique molecular identifiers (UMIs) associate distinct sequences with every DNA or RNA molecule and can be counted after amplification to quantify molecules in the original sample. Using UMIs, the authors obtain a digital karyotype of an individual with Down's syndrome and quantify mRNA in Drosophila melanogaster cells.

Sensitive and specific single-molecule sequencing of 5-hydroxymethylcytosine pp75 - 77

Chun-Xiao Song, Tyson A Clark, Xing-Yu Lu, Andrey Kislyuk, Qing Dai, Stephen W Turner, Chuan He & Jonas Korlach


The DNA modification 5-hydroxymethylcytosine has recently been implicated in several biological processes. Enrichment by selective chemical labeling in combination with single-molecule, real-time sequencing provides sensitive detection of this epigenetic mark in genomic DNA at base-pair resolution.

Decoding cell lineage from acquired mutations using arbitrary deep sequencing pp78 - 80

Cheryl A Carlson, Arnold Kas, Robert Kirkwood, Laura E Hays, Bradley D Preston, Stephen J Salipante & Marshall S Horwitz


Mutations at arbitrarily sampled genomic positions are identified using next-generation sequencing and are used to infer the lineage of DNA damage–prone 'mutator' mouse cells in culture.

Controlled gene expression in primary Lgr5 organoid cultures pp81 - 83

Bon-Kyoung Koo, Daniel E Stange, Toshiro Sato, Wouter Karthaus, Henner F Farin, Meritxell Huch, Johan H van Es & Hans Clevers


The controlled overexpression or knockdown of gene expression in primary organoid cultures of mouse endodermal epithelia is described. This should enable ex vivo studies of mammalian gene function.



Global profiling of dynamic protein palmitoylation pp84 - 89

Brent R Martin, Chu Wang, Alexander Adibekian, Sarah E Tully & Benjamin F Cravatt


A quantitative proteomics approach to characterize protein palmitoylation dynamics on a global scale in cells, as well as to identify enzymes responsible for the regulation of palmitoylation, is described.

Optical recording of action potentials in mammalian neurons using a microbial rhodopsin pp90 - 95

Joel M Kralj, Adam D Douglass, Daniel R Hochbaum, Dougal Maclaurin & Adam E Cohen


The microbial rhodopsin protein, Archaerhodopsin 3, can function as a rapid and highly sensitive genetically encoded voltage indicator in mammalian cells that is capable of detecting single action potentials with a signal-to-noise ratio greater than 10. A mutant lacking proton pumping displays greater sensitivity but a slowed response.

mGRASP enables mapping mammalian synaptic connectivity with light microscopy pp96 - 102

Jinhyun Kim, Ting Zhao, Ronald S Petralia, Yang Yu, Hanchuan Peng, Eugene Myers & Jeffrey C Magee


In this paper, the authors report GFP reconstitution across synaptic partners (GRASP) adapted for synapse visualization in the mammalian brain.

High-efficiency counterselection recombineering for site-directed mutagenesis in bacterial artificial chromosomes pp103 - 109

Alexander W Bird, Axel Erler, Jun Fu, Jean-Karim Hériché, Marcello Maresca, Youming Zhang, Anthony A Hyman & A Francis Stewart


Site-directed seamless modification of bacterial artificial chromosomes is enhanced more than tenfold in efficiency by improving the counterselection step. A set of plasmids and oligonucleotide design software also make this E. coli recombineering approach markedly faster and easier.




Erratum: Reply to “More on color blindness” p110

Bang Wong