Monthly methods overview

Here is a selection of interesting new methods published in the literature as curated by Nature Methods editors. You can find our top picks this month in our September issue, which has just come out.
Published in Protocols & Methods
Monthly methods overview

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The Nature Methods editorial team gets together each month to pick interesting new methods developments that are published in other journals for the Research Highlights section. Unfortunately, we can't highlight all the great methods being developed. Below are a few exciting papers that we couldn't feature in our September issue.


Basanta, B. et al. An enumerative algorithm for de novo design of proteins with diverse pocket structures. PNAS (2020).

A Rosetta-based approach enables enumerative de novo design of protein structures with the same backbone conformation but diverse pocket geometries.  The authors design ~8000 nuclear transport factor 2 (NFT2) family-like proteins, experimentally test them for stability, and characterize several biophysically. Crystal structures obtained for 5 of these designs highlight the diversity achievable by their approach.

Yu, F. et al. Identification of modified peptides using localization-aware open search. Nat. Commun. 11, 4065 (2020).

A strategy for database searching allows the identification of post-translationally or chemically modified peptides in mass spectrometry-based proteomics experiments. It is implemented in MSFragger2.0. The approach uses a shifted ion index and localization-aware open search for fragment ion matching and scoring.

Carrasco-López, C. et al. Development of light-responsive protein binding in the monobody non-immunoglobulin scaffold. Nat. Commun. 11, 4045 (2020)

A light-controlled monobody (OptoMB) that targets an SH2 domain allows for targeted and reversible optogenetic control of binding  in vitro and in cells. The OptoMB was used for rapid and efficient purification of SH2-tagged proteins from crude E. coli extract.

Gil, A.A. et al. Optogenetic control of protein binding using light-switchable nanobodies. Nat. Commun. 11, 4044 (2020).

Opto-nanobodies (OptoNBs) are chimeric photoswitchable proteins whose ability to bind target molecules can be enhanced or inhibited by treatment with blue light. OptoNB binding is reversible, and they can be used in a broad range of applications in vitro and in cells for controllable regulation of a variety of targets. 


Su, J.-H. et al. Genome-Scale Imaging of the 3D Organization and Transcriptional Activity of Chromatin. Cell 182, 1-19 (2020).

The 3D organization of the genome and its relationship to gene expression is explored using a high-throughput platform that probes 1,000 genomic loci along with the transcription of more than 1,000 genes in thousands of single cells.

Bergen, V. et al. Generalizing RNA velocity to transient cell states through dynamical modeling. Nat. Biotechnol. (2020).

RNA velocity reveals rich information about single-cell gene expression dynamics based on the ratio of spliced and unspliced mRNAs. Bergen et al. develop a computational tool called scVelo that generalizes RNA velocity to transient systems and systems with heterogeneous kinetics. Using likelihood-based dynamical models, scVelo leads to findings about gene regulation and cell fate decisions in neurogenesis and pancreatic endocrinogenesis.

Li, X. et al. Dynamic incorporation of multiple in silico functional annotations empowers rare variant association analysis of large whole-genome sequencing studies at scale. Nat. Genet. 52, 969–983 (2020).

Whole-genome/exome sequencing unveils large numbers of rare variants, whose contributions to phenotypes and diseases are often hard to ascertain statistically. Li et al. present STARR, a rare-variant association test method that leverages functional annotations using dynamic weighting. Application of STARR in the large-scale TOPMed dataset finds new associations with lipid traits.

Lapinaite, A. et al. DNA capture by a CRISPR-Cas9–guided adenine base editor. Science 369, 566–571 (2020).

Base editors have become a promising tool in the gene-editing toolbox. This paper applies single-particle cryo-EM to characterize the structure of the ABE8e complex, which contains Cas9, sgRNA and DNA target-strand, and reveals the underlying molecular basis for the different catalytic rates and base-editing outcomes offered by ABE7.10 and ABE8e variants.

You, Q. et al. Direct DNA crosslinking with CAP-C uncovers transcription-dependent chromatin organization at high resolution. Nat. Biotechnol. (2020).

3C or Hi-C based approaches require crosslinking proteins with genomic DNA for probing proximity between genomic loci. However, the presence of DNA-bound proteins might mask enzymatic digestion sites and reduce the resolution of chromatin contact maps. To reduce the background noise, this paper describes chemical crosslinking-assisted proximity capture (CAP-C), which uses synthetic crosslinker PAMAM dendrimers to crosslink DNA and capture proximal DNA loci for generating contact maps with sub-kilobase resolution. 

Lareau, C.A. et al. Massively parallel single-cell mitochondrial DNA genotyping and chromatin profiling. Nat. Biotechnol. (2020).

This paper describes a droplet-based assay - mitochondrial single-cell assay for transposase-accessible chromatin with sequencing (mtscATAC-seq) - that concomitantly identifies mitochondrial DNA variants and profiles transposase-accessible mtDNA in single cells. 


Di Antonio, M. et al. Single-molecule visualization of DNA G-quadruplex formation in live cells. Nat. Chem. 12, 832–837 (2020). 

A probe that specifically binds G4 quadruplexes in DNA (SiR-PyPDS) enables single-molecule and real-time detection of individual G4 structures in living cells without globally perturbing G4 formation and dynamics.

Bazhin, A.A. et al. A bioluminescent probe for longitudinal monitoring of mitochondrial membrane potential. Nat. Chem. Biol. (2020).

Mitochondrial membrane potential can now be measured in vivo with a bioluminescent probe. Mitochondria-activatable luciferin (MAL) releases active luciferin in relation to mitochondrial membrane potential to allow for non-invasive and longitudinal monitoring.


Damo, M. et al. Inducible de novo expression of neoantigens in tumor cells and mice. Nat. Biotechnol. (2020).

Damo et al. have developed an inversion-induced joined neoantigen (NINJA) mouse model to induce de novo expression of neoantigens. A tightly regulated process involving gene induction based on delivery of Cre- recombinase, Tamoxifen and Doxycycline ensures tissue-specific expression of neoantigens to study endogenous antigen-specific T cell responses.

Salzer, B. et al. Engineering AvidCARs for combinatorial antigen recognition and reversible control of CAR function. Nat. Commun. (2020).

Salzer et al. present a logic gate-based platform to control CAR T cell function. By integrating an ON switch within the CAR molecule, low-affinity CARs can be programmed to dimerize in the presence of a soluble factor or upon recognizing two different antigens, thus increasing the antigen sensitivity of the anti-tumor response.


Ichino, N. et al. Building the vertebrate codex using the gene breaking protein trap library. Elife 9:e54572 (2020).

Ichino et al. present a collection of 1200 zebrafish lines harboring protein traps that were generated with the gene-breaking transposon technology. The inserted transposons typically result in mRNA knockdown in homozygous animals as well as the expression of an mRFP reporter under the control of the targeted gene's promoter in both homozygous and heterozygous animals. The zebrafish lines have been cryopreserved and are available from a stock center.


Amunts, K. et al. Julich-Brain: A 3D probabilistic atlas of the human brain's cytoarchitecture. Science 369, 988992 (2020).

The Julich-Brain is a 3D cytoarchitectonic atlas of the human brain, which is based on histological sections from 23 brains and annotations from 41 projects focussing on different brain areas. The atlas takes individual variability into account, is dynamic to enable incorporation of new data and can link to other atlases and resources. The atlas is available at

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