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CRISPR–Cas9 has truly democratized genome editing. In this Perspective, Jin-Soo Kim discusses CRISPR–Cas9 genome editing in the context of earlier innovations using meganucleases, ZFNs and TALENs, which paved the way for the ongoing CRISPR–Cas revolution.
iPSC technology has radically advanced our understanding of mammalian development. In this Perspective, Belmonte and Li discuss breakthroughs in iPSC technology over the past decade and share their predictions for the future of iPSCs in developmental biology and regenerative medicine.
Targeted DamID (TaDa) extends DamID to enable cell-type-specific profiling of genome-wide protein binding. Transcription factor binding sites, RNA Pol II occupancy and chromatin states can be studied to provide insights into cell-fate specification.
This protocol enables high-throughput sequencing analysis of the full-length immunoglobulin repertoires in human and mouse using unique molecular identifiers (UMIs) and asymmetric paired-end sequencing.
This protocol describes how to perform whole-mount immunostaining and imaging on adult mouse small-intestinal villi. All gut cell types can be seen at high resolution and in 3D without the need for image reconstruction.
SUMO (small ubiquitin-like modifier) family proteins are post-translationally attached to lysine residues in a variety of proteins. This protocol describes a method for mass-spectrometry-based identification of SUMOylated lysines in mammalian cells.
Pertea et al. describe a protocol to analyze RNA-seq data using HISAT, StringTie and Ballgown (the ‘new Tuxedo’ package). The protocol can be used for assembly of transcripts, quantification of gene expression levels and differential expression analysis.
This is a relatively simple protocol for the routine culture of P. falciparum gametocytes in vitro. The functional viability of gametocytes is assessed, and they can then be used to evaluate transmission-blocking activity of drugs.
This protocol describes stepwise differentiation of human pluripotent stem cells into 3D kidney organoids that contain segmented nephrons connected to collecting ducts, which are surrounded by renal interstitial cells and an endothelial network.
This protocol describes a high-content microscopy approach to quantifying mitochondrial morphofunction in living cells. After staining with fluorescent reporters, images are automatically acquired, processed and analyzed to extract 44 descriptors.
This protocol describes a simple and effective method for live imaging of axonal transport in adult neurons. It makes use of the translucent Drosophila wing and spinning-disk microscopy to obtain high-resolution movies of axonal cargoes in transit.
This protocol describes the long-term culture of liver and pancreas 3D organoids from human and mouse, and differentiation of liver organoids in vitro and in vivo. Methodology for genetic manipulation of these self-renewing organoids is also detailed.
Shigemitsu et al. describe the preparation of supramolecular hydrogelators that undergo redox-responsive gel degradation. Encapsulation of a variety of enzymes in the gels allows for degradation in response to various biomolecules.
Cell Painting is a high-content screening assay that uses multiplexed fluorescent dyes for image-based profiling of ∼1,500 morphological features. Image analysis with CellProfiler automatically identifies and extracts data from individual cells.