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The 3′ region extraction and deep sequencing (3′READS) method accurately identifies cleavage and polyadenylation sites, avoiding common artifacts and detecting sites in A-rich contexts. It was used to greatly expand the number of characterized sites in the mouse genome, including those in long noncoding RNAs.
A collection of simulation tools and workflow for single-molecule Förster resonance energy transfer (smFRET) allows highly quantitative structural modeling. This hybrid approach yields a model of reverse-transcriptase binding to DNA at sub-angstrom accuracy when benchmarked against a crystal structure and can resolve a flexible single-stranded template overhang.
The structure of the membrane anchor domain of the bacterial autotransporter YadA is solved by a solid-state NMR spectroscopy approach using a uniformly 13C- and 15N-labeled microcrystalline sample.
Cells are dosed with magnetic nanoparticles and patterned onto micromagnetic substrates, enabling the application of controlled and variable mechanical force to tens of thousands of cells.
The probe selection for imputation (PSI) approach accurately imputes global gene expression profiles from a small subset of probes that it chooses based on a training set of full profiles, allowing many more combinatorial experiments to be performed given the same resources.
The Strand-seq method independently sequences each parental strand of template DNA from single proliferating cells. It can be used to detect sister chromatid exchange and other chromosomal abnormalities at high resolution and to correct contig misorientations in genome assemblies, with potential for strand-inheritance and haplotyping studies.
A synthetic peptide library in conjunction with liquid chromatography–tandem mass spectrometry identifies the specificities of endo- and exopeptidases without requiring enrichment of substrates or products.
A genetic tool that converts bacterial regulators of translational initiation into regulators of transcriptional elongation is described. This adaptor is used to engineer several transcriptional attenuators and activators that can be predictably assembled into higher-order gene regulatory functions.
A G protein–coupled receptor (GPCR) signaling assay based on ectodomain shedding of a membrane-bound proform of alkaline phosphatase-tagged TGFα provides a platform for studying poorly characterized Gα12/13-coupled GPCRs. The assay allowed identification of three orphan GPCRs as Gα12/13-coupled lysophosphatidylserine receptors.
Development of the bright green and red fluorescent proteins, Clover and mRuby2, creates a fluorescence resonance energy transfer (FRET) pair with the highest Förster radius among existing ratiometric FRET pairs. Substitution of this pair for current FRET pairs in several existing sensors reliably and substantially improves sensor performance.
A dual-trap force-clamp configuration is used to apply a constant load between a binding protein and a single intermittently interacting biological polymer. This allows high-resolution measurements of short-lived molecular complexes and reveals previously undetected complex regulation of the myosin working stroke.
This paper reports genetic manipulation of the malaria parasite Plasmodium falciparum with zinc-finger nucleases. It demonstrates gene disruption as well as replacement and site-specific editing of both an integrated reporter and an endogenous gene.
A method is presented to model cell proliferation from tracked automated time-lapse imaging of cell populations. The resulting fractional proliferation graphs permit visualization of dynamic changes in the dividing and nondividing subsets of the population.
A flexible, environmentally controlled experimental setup for the study of terrestrial animal dispersal is reported. Its unprecedented scale should enable studies in spatial ecology and permit tests of conservation strategies in the face of environmental change.
Primary rat oligodendocytes were cultured in the presence of electron-spun nanofibers of varying sizes as a model to study myelination processes in the mammalian central nervous system. The authors study the role of fiber diameter on the initiation of concentric wrapping by oligodendrocytes.
Two large-scale resources for studying microRNA function are presented: one is a library of fluorescent sensors with a corresponding assay for global profiling of microRNA activity in different cell types; the other is a decoy library for suppressing microRNA activity individually or in pooled loss-of-function screens.