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An integrated, miniature (1.9 g) fluorescence microscope containing light source, optics and sensor allows high-speed, wide field of view imaging of calcium spiking in hundreds of neurons in freely moving mice. The mass-producible portable microscope is also useful for a variety of fluorescence assays for which size, cost and portability can be concerns.
This paper reports transgenesis by genetic modification of gametes in the domestic cat. The approach is used to generate transgenic cats expressing a virus restriction factor from rhesus macaque.
An algorithm for linear mixed models substantially reduces memory usage and run time for genome-wide association studies. The improved algorithm scales linearly in cohort size, allowing the application of these models to much larger samples.
Reprogramming to induced pluripotency of cells from the endangered silver-maned drill and the northern white rhinoceros is reported. Induced pluripotent stem cells from endangered species may prove useful for species preservation in the future.
Expression profiles of several hundred microRNAs in the blood of individuals with disease, including autoimmune disease, cancers, cardiovascular disease and chronic inflammatory disease are reported.
Destabilized mutants of firefly luciferase are characterized as sensors for protein homeostasis (proteostasis). Their use as tools for comparisons of proteostasis capacity is demonstrated in cells and in Caenorhabditis elegans.
A method for performing quantitative proteomics experiments in Caenorhabditis elegans using stable-isotope labeling of lysine is described. The method can be coupled with RNA interference to examine global effects on the proteome. Also in this issue, Larance et al. describe a very similar method.
A method for performing quantitative proteomics experiments in Caenorhabditis elegans using stable-isotope labeling with amino acids in cell culture (SILAC) is described. The authors used the method to identify heat shock–responsive proteins in worms. Also in this issue, Fredens et al. describe a very similar method.
Presented is an experimental analysis of the stability of transgene expression, the perturbation of endogenous expression and the perturbation of epigenetic organization upon site-directed delivery of transgenes to the CCR5 and AAVS1 loci in human cells. It provides guidelines for optimal cassette design for stable and nonperturbative gene transfer.
An in vitro method for examining cleavage patterns of zinc-finger nucleases (ZFNs) identifies previously unknown off-target cleavage sites. Some of the sites are present in the human genome and show evidence for ZFN-induced cleavage in cultured human cell lines.
A collection of functionally characterized BAC transgenic mouse lines in which the channelrhodopsin-2 H134R variant is specifically and stably expressed in GABAergic, cholinergic, serotonergic or parvalbumin-positive neurons is reported.
Described is a randomly inserting transposon that can be swapped for gene traps, coding insertions or protein tag genes, thus expanding the toolkit for Drosophila melanogaster genome engineering.
A fluorescent protein that can be photoswitched with visible light from orange to far-red is presented. The photoconverted form has the most red-shifted excitation peak of all GFP-like fluorescent proteins to date and should be useful for many imaging applications.
SourceTracker finds the proportion and origin of contaminants in a given sample. Its database will prove useful in screening of metagenomic datasets for contaminants.
Single-stranded oligonucleotides are used as donor templates for zinc-finger nucleases to create targeted genomic deletions and mutations in mammalian cell lines.
Two-photon excitation in scanned light-sheet microscopy allows deeper imaging than the one-photon implementation and faster three-dimensional imaging of organism development with no evidence of phototoxicity compared to conventional two-photon point scanning microscopy.
Quantitative, large-scale in vivo phosphoproteomics analyses are made possible with a form of spike-in stable-isotope labeling with amino acids in cell culture (SILAC), in which SILAC-labeled cell lines act as an internal standard for mass spectrometry–based tissue phosphoproteome analysis.
A fluorescent reporter, named traffic light, reads out whether repair of a DNA break occurs by nonhomologous end-joining or by homologous recombination. It should enable the identification of factors that affect repair pathway choice and thus improved approaches for genome engineering.
Ubiquitin, an important post-translational modification that regulates a variety of biological processes is found in free and conjugated (monoubiquitin and polyubiquitin) forms in the cell. A method for precisely measuring these cellular pools using protein standard absolute quantification mass spectrometry is described; the approach should yield insights into ubiquitin signaling.
This digital PCR device arrays samples into one million small-volume reactors, achieving a dynamic range of 107, measurement precision better than 1% and the ability to detect single-nucleotide variants present at less than 1:100,000.