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    Protocols in Current Issue
    A Protocol to Map the Spatial Proteome Using HyperLOPIT in Saccharomyces cerevisiae
    Authors:  Daniel J. H. Nightingale, Kathryn S. Lilley and Stephen G. Oliver, date: 07/20/2019, view: 29, Q&A: 0
    [Abstract] The correct subcellular localization of proteins is vital for cellular function and the study of this process at the systems level will therefore enrich our understanding of the roles of proteins within the cell. Multiple methods are available for the study of protein subcellular localization, including fluorescence microscopy, organelle ...
    Preparation of RNA 3’ End Sequencing Libraries of Total and 4-thiouracil Labeled RNA for Simultaneous Measurement of Transcription, RNA Synthesis and Decay in S. cerevisiae
    Authors:  Manfred Schmid, Agnieszka Tudek and Torben Heick Jensen, date: 03/20/2019, view: 774, Q&A: 0
    [Abstract] Cellular RNA levels are determined by the rates of RNA transcription from the gene template and subsequent RNA stability. Knowledge about both transcription and RNA decay is, therefore, necessary to interpret RNA levels and gene expression, especially during cellular processes where these parameters change. Numerous experimental strategies have ...
    Assessing Yeast Cell Survival Following Hydrogen Peroxide Exposure
    Authors:  Khoa Tran and Erin M. Green, date: 01/20/2019, view: 1787, Q&A: 0
    [Abstract] In the presence of oxidative stress, cellular defense systems that can detoxify reactive oxygen species are activated through multiple signaling cascades and transcriptional reprogramming. The budding yeast Saccharomyces cerevisiae has served as an excellent model for genetically-identifying factors important for the response to oxidative ...
    An in vitro Microscopy-based Assay for Microtubule-binding and Microtubule-crosslinking by Budding Yeast Microtubule-associated Protein
    Authors:  Yili Zhu, Weimin Tan and Wei-Lih Lee, date: 12/05/2018, view: 1444, Q&A: 0
    [Abstract] In this protocol, we describe a simple microscopy-based method to assess the interaction of a microtubule-associated protein (MAP) with microtubules. The interaction between MAP and microtubules is typically assessed by a co-sedimentation assay, which measures the amount of MAP that co-pellets with microtubules by centrifugation, followed by ...
    Separation and Visualization of Low Abundant Ubiquitylated Forms
    Authors:  Ramona Schuster, Tânia Simões, Fabian den Brave and Mafalda Escobar-Henriques, date: 11/20/2018, view: 1266, Q&A: 0
    [Abstract] In this protocol we describe the separation and visualization of ubiquitylated forms of the yeast mitofusin Fzo1 by Western blot. To this aim, we express HA-tagged Fzo1 in Saccharomyces cerevisiae, break the cells to extract a membrane-enriched fraction, solubilize the membranes using detergent and then specifically immunoprecipitate the ...
    Preparation of Sequencing RNA Libraries through Chemical Cross-linking Coupled to Affinity Purification (cCLAP) in Saccharomyces cerevisiae
    Authors:  Congwei Wang, Julie Weidner and Anne Spang, date: 10/05/2018, view: 1837, Q&A: 0
    [Abstract] Ribonucleoprotein particles (mRNPs) are complexes consisting of mRNAs and RNA-binding proteins (RBPs) which control mRNA transcription localization, turnover, and translation. Some mRNAs within the mRNPs have been shown to undergo degradation or storage. Those transcripts can lack general mRNA elements, like the poly(A) tail or 5’ cap structure, ...
    Extracting and Integrating Protein Localization Changes from Multiple Image Screens of Yeast Cells
    Authors:  Alex X Lu, Louis-Francois Handfield and Alan M Moses, date: 09/20/2018, view: 1271, Q&A: 0
    [Abstract] The evaluation of protein localization changes in cells under diverse chemical and genetic perturbations is now possible due to the increasing quantity of screens that systematically image thousands of proteins in an organism. Integrating information from different screens provides valuable contextual information about the protein function. For ...
    Detection of Internal Matrix Targeting Signal-like Sequences (iMTS-Ls) in Mitochondrial Precursor Proteins Using the TargetP Prediction Tool
    Authors:  Felix Boos, Timo Mühlhaus and Johannes M. Herrmann, date: 09/05/2018, view: 1520, Q&A: 0
    [Abstract] Mitochondria contain hundreds of proteins which are encoded by the nuclear genome and synthesized in the cytosol from where they are imported into the organelle. Sorting signals encoded in the primary and secondary sequence of these proteins mediate the recognition of newly synthesized precursor proteins and their subsequent translocation through ...
    A Procedure for Precise Determination of Glutathione Produced by Saccharomyces cerevisiae
    Authors:  Jyumpei Kobayashi, Daisuke Sasaki and Akihiko Kondo, date: 06/20/2018, view: 2388, Q&A: 0
    [Abstract] In bioproduction, yields of products must be calculated precisely for accurate evaluation of various fermentation conditions. To evaluate productivity of microorganisms, product amounts per unit of medium volume (e.g., mg-product/L-broth), and/or product amounts per unit of a microorganism amount (e.g., mg-product/mg-dry cell ...
    Dual-probe RNA FRET-FISH in Yeast
    Authors:  Gable M. Wadsworth, Rasesh Y. Parikh and Harold D. Kim, date: 06/05/2018, view: 2452, Q&A: 0
    [Abstract] mRNA Fluorescence In Situ Hybridization (FISH) is a technique commonly used to profile the distribution of transcripts in cells. When combined with the common single molecule technique Fluorescence Resonance Energy Transfer (FRET), FISH can also be used to profile the co-expression of nearby sequences in the transcript to measure ...



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