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A simple automated system to simultaneously measure growth rate and lag time of large numbers of bacteria permits the identification of subsets in a heterogenous population.
Micropost arrays can be used to modulate substrate rigidity independently of other substrate properties, permitting the study of the effects of rigidity on cell function.
A platform for rapid and automated imaging and laser manipulation of zebrafish larvae is presented. It should permit large-scale chemical and genetic screens in this vertebrate organism.
On-flowcell capture and reverse transcription followed by single-molecule cDNA sequencing provides reproducible digital gene expression results from as few as 1,000 cells.
Single integration of a target gene expressing MS2-binding stem loops allows the real time quantification of transcriptional bursts, promoter firings and cell cycle–dependent transcription rates.
Improving the protocols for chromatin immunoprecipitation and library construction for the Illumina Genome Analyzer allows for chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq) experiments on input samples as small as 10,000 cells and yields information on bivalent chromatin domains in hematopoietic progenitor cells.
A monomeric fluorescent protein that can be irreversibly photoswitched from green to red form, both of which can be reversibly photoactivated, is reported. It is applied to pulse-chase experiments in which dynamic structures in live cells are imaged with superresolution using photoactivation localization microscopy (PALM).
A genetically encoded ratiometric biosensor not based on fluorescence resonance energy transfer (FRET), ClopHensor, allows concurrent measurement of intracellular pH and chloride by providing an internal control for pH-dependent fluorescence changes. Measurements of chloride levels in acidic large dense core vesicles showed high concentrations of chloride.
Light-sensitive LOV domains show much promise for engineering proteins with photoswitchable activity. The dynamic range of a LOV domain is now substantially improved by the introduction of beneficial mutations predicted by an analytical model of photoswitching. The approach should prove useful to improve the function of multiple LOV-based switches.
A method to analyze the sequence of C-terminal peptides using a combination of a specific enrichment approach and mass spectrometry is described, allowing the study of C-terminal proteolytic processing on a global scale. Also in this issue, Van Damme et al. describe a related method for simultaneous C- and N-terminal peptide analysis.
A method to simultaneously analyze C- and N-terminal peptides using a combination of strong cation exchange, diagonal chromatography and mass spectrometry is described, allowing the screening of protease substrates on a global scale. Also in this issue, Schilling et al. describe a related method for analyzing the sequence of C-terminal peptides.
The misfolded form of the prion protein, PrPSc, can be quantified in a variety of tissues and fluids using a quantitative version of the popular protein misfolding cyclic amplification (PMCA) assay.
Cultured rat spermatogonia, genetically modified via a transposon-based gene trap, are inserted into the testes of sterile rats and give rise to mutant progeny with many different genotypes.
The activity of zinc-finger nucleases in mammalian cells is enhanced by transient incubation of the cells at low temperatures. Incorporation of this simple step should improve the efficiency of these tools for genome manipulation.
This computational process evaluates gene models in prokaryotic genomes, independently of the gene finder used, and reports anomalies that can be used to improve the quality of gene models through manual curation.
Targeted deletion of genes within 25 kb of Mos1 transposons in the C. elegans genome is demonstrated. This should enable single-gene deletions of more than 8,000 genes for which deletions are currently not available in this organism.
Traptavidin, a streptavidin mutant with about tenfold lower 'off' rate for biotin than streptavidin itself, has increased mechanical strength and thermostability. It should find use in a diversity of applications in which the dissociation of streptavidin can be a limitation.
A microfluidic device containing a suspended microchannel resonator capable of measuring the mass of microscopic objects with femtogram resolution allows determination of bacteria, yeast and mammalian cell growth rates in less than one cell cycle by repeated measurement of the buoyant mass of single growing cells.
