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HHblits is a protein sequence search tool that works by iterative pairwise comparison of profile hidden Markov models. It outperforms existing tools in terms of speed, sensitivity and alignment quality.
The Splitread algorithm uses a split-read strategy to detect structural variants and small insertions and deletions (indels) in whole-exome and whole-genome sequence datasets at high sensitivity. It maps the breakpoints at single-base-pair resolution, even in low-complexity regions, and can detect novel processed pseudogenes.
An efficient haplotype-estimation algorithm that features linear complexity allows the rapid and accurate phasing of diploid genomes from trios, duos and unrelated samples.
The controlled overexpression or knockdown of gene expression in primary organoid cultures of mouse endodermal epithelia is described. This should enable ex vivo studies of mammalian gene function.
Mutations at arbitrarily sampled genomic positions are identified using next-generation sequencing and are used to infer the lineage of DNA damage–prone 'mutator' mouse cells in culture.
The DNA modification 5-hydroxymethylcytosine has recently been implicated in several biological processes. Enrichment by selective chemical labeling in combination with single-molecule, real-time sequencing provides sensitive detection of this epigenetic mark in genomic DNA at base-pair resolution.
Unique molecular identifiers (UMIs) associate distinct sequences with every DNA or RNA molecule and can be counted after amplification to quantify molecules in the original sample. Using UMIs, the authors obtain a digital karyotype of an individual with Down's syndrome and quantify mRNA in Drosophila melanogaster cells.
Conjugation of triplet-state quenchers to the small organic cyanine fluorophore, Cy5, increases photostability without affecting its spectral characteristics. This allows longer fluorescence imaging with a concomitant reduction in blinking both in vitro and in living cells.
Presented is a study of gene regulation during development using a combination of chromatin immunoprecipitation and high-throughput sequencing (ChIP-seq) and directed differentiation of mouse embryonic stem cells inducibly expressing epitope-tagged transcription factors.
An empirical approach for identifying optimal proteotypic peptides and fragmentation patterns from in vitro–synthesized proteins, for targeted proteomics applications, is described.
An almost-complete, sequence-verified collection of Arabidopsis thaliana root stele transcription factors is reported. The authors use it in the enhanced yeast-one hybrid (eY1H) assay to map gene regulatory interactions in the plant. Also in this issue, Reece-Hoyes et al. describe the eY1H pipeline.
A sequence-verified collection of human transcription factors is reported. The authors used it in the enhanced yeast-one hybrid (eY1H) assay to map human gene regulatory networks. Also in this issue, Reece-Hoyes et al. describe the eY1H pipeline.
A fluorescent molecular tension sensor for spatially and temporally mapping the mechanical strain exerted by cell-surface receptors in living cells is described.
A human embryonic stem cell line expressing a fluorescent reporter of cardiac differentiation is described. The authors use this tool to optimize differentiation methods and to identify cell-surface markers in the cardiac lineage.
Live-cell volumetric super-resolution imaging with 120-nm lateral and 360-nm axial resolution using structured-illumination microscopy at speeds of up to 5 s per cell volume over >50 time points captures fine cellular dynamics using only low illumination intensities.
The combination of light-sheet microscopy and localization-based super-resolution imaging allows deep subdiffraction resolution imaging in thick scattering specimens as demonstrated by three-dimensional super-resolution imaging of proteins in live 150-μm-diameter cell spheroids.
A simple episomal fluorescent reporter for flow cytometric enrichment of cells with zinc-finger nuclease– or TALE nuclease–induced genomic modifications is described.
A computational approach for analysis of gene expression in heterogeneous samples of varying composition is presented. The authors used it to study expression in brain samples from humans with Huntington's disease.
A triple-stage mass spectrometry (MS3)-based method is used to remove ratio interference, resulting in accurate, large-scale, multiplexed quantitative proteomics measurements using isobaric labeling. Also in this issue, Wenger et al. provide a different solution to the same problem.
A mass spectrometry instrument control method—QuantMode—allows accurate, large-scale, multiplexed quantitative proteomics measurements using isobaric tagging by removing the problem of precursor interference. Also in this issue, Ting et al. provide a different solution to the same problem.
