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Correia-Melo et al. describe a protocol to generate and maintain mitochondria-depleted mammalian cell lines. These cells can be used to investigate the role of mitochondria in various cellular processes such as cell death and senescence.
This protocol describes how to differentiate human pluripotent stem cells into nephron progenitor cells with subsequent generation of 2D and 3D kidney organoids.
This protocol describes surgical techniques for long-term intrathecal drug administration to rodent CNS tissue and cerebrospinal fluid collection for monitoring of drug concentration and distribution.
Correlative light and electron microscopy (CLEM) promises new insight into cellular processes by combining spatiotemporal information with high-resolution structural information. This protocol describes cryo-CLEM of virus-infected mammalian cells.
This protocol describes the diastereoselective approach for the synthesis of complex, acyclic molecular architectures possessing two stereogenic centers in a 1,4 relationship based on a zirconium-promenade reaction using the Negishi reagent.
This protocol uses a doxycycline-inducible Cas9 transgene carried on a piggyBac transposon for robust, highly efficient and scarless genome editing of human iPSCs, enabling a parental line to be engineered to harbor many separate mutations.
Zilionis et al. describe a protocol for the inDrops platform, which is a high-throughput droplet microfluidics system that allows for single-cell barcoding of over 15,000 cells in 1 h.
Different concentrations of activin A and BMP4 are used to polarize stem cells into mesodermal subtypes that reflect mid-primitive-streak cardiogenic mesoderm and posterior-primitive-streak hemogenic mesoderm.
Standard methods for antibody staining of Drosophila larvae have worked poorly or involved dissection. The Doe laboratory describes a reliable protocol for immunofluorescent antibody staining of intact Drosophila larvae that works well for all larval stages.
Comprehensive analysis of membrane protein interactions using COPIT includes information for transient and weak interactions. This mass spectrometry–based method has optimized steps for removing contaminants and a tailored data analysis.
Fang et al. describe a computational protocol to accurately call indels from whole-genome and whole-exome sequencing data using Scalpel. Important issues for indel identification, such as short repeat regions and varying sequencing coverage, are discussed.
This protocol from Rodrigues et al. describes a cell-free in vitro system to examine the machinery that mediates peroxisomal protein import. The system allows the user to block components of the machinery at virtually any step.
This protocol describes a nondestructive FRET-based assay for measuring protein interactions in cultured cells. The correlation between FRET efficiencies and relative protein concentration can be used to determine relative binding affinities.
White et al. present a protocol for ARQiv-HTS, a high-throughput reporter-based screening platform that can be used to examine the effects of chemical compound libraries on zebrafish in vivo.
Garcia et al. describe a protocol for mapping-by-sequencing in Tomato. After selecting EMS mutants displaying a phenotype of interest, mutants are back-crossed and causal mutations are identified in the F2 population using sequencing-based approaches.
Primed conversion of photoconvertible fluorescent proteins enables targeted labeling in vivo with high axial resolution. This Protocol describes how to modify a confocal microscope to perform confined primed conversion of Dendra2 in living organisms.