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Live Intravital Intestine with Blood Flow Visualization in Neonatal Mice Using Two-photon Laser Scanning Microscopy

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Molecular and Phenotypic Characterization Following RNAi Mediated Knockdown in Drosophila

Authors: Saurabh Jayesh Kumar Mehta, Pradyumna A. Joshi and Ram Kumar Mishra, date: 02/20/2021, view: 667, Q&A: 0

[Abstract]

Loss of function studies shed significant light on the involvement of a gene or gene product in different cellular processes. Short hairpin RNA (shRNA) mediated RNA interference (RNAi) is a classical yet straightforward technique frequently used to knock down a gene for assessing its function. Similar perturbations in gene expression can be

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DigiTAG–a RNA Sequencing Approach to Analyze Transcriptomes of Rare Cell Populations in Drosophila melanogaster

Authors: Lisa Landskron, Francois Bonnay, Thomas R. Burkard and Jürgen A. Knoblich, date: 11/05/2020, view: 927, Q&A: 0

[Abstract]

Cell-type specific transcriptional programs underlie the development and maintenance of organs. Not only distinct cell types within a tissue, even cells with supposedly identical cell fates show a high degree of transcriptional heterogeneity. Inevitable, low cell numbers are a major hurdle to study transcriptomes of pure cell populations. Here we

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Production and Bioassay of a Diffusible Factor that Induces Gametophyte-to-Sporophyte Developmental Reprogramming in the Brown Alga Ectocarpus

Authors: Haiqin Yao, Delphine Scornet, Yacine Badis, Akira F. Peters, Murielle Jam, Cecile Hervé, Philippe Potin, Susana M. Coelho and J. Mark Cock, date: 09/20/2020, view: 1514, Q&A: 0

[Abstract] The brown alga Ectocarpus has a haploid-diploid life cycle that involves alternation between two multicellular generations, the sporophyte and the gametophyte. Life cycle generation is not determined by ploidy but by a genetic system that includes two different three amino acid loop extension homeodomain transcription factors called ...

In vitro Induction and Detection of Acrosomal Exocytosis in Human Spermatozoa

Authors: Shenae L. Cafe, Amanda L. Anderson and Brett Nixon, date: 07/20/2020, view: 959, Q&A: 0

[Abstract] The acrosome reaction is a highly regulated exocytotic event that primes spermatozoa for successful fertilization. Upon induction, acrosomal exocytosis proceeds via a wave of vesiculation that radiates across the sperm head, destabilizing the acrosomal vesicle and resulting in the release of the acrosomal contents. Having shed their acrosome, ...

Organotypic Slice Culture of the Embryonic Mouse Brain

Authors: James M. Clegg and Thomas Pratt, date: 07/05/2020, view: 1094, Q&A: 0

[Abstract] Organotypic slice culture is a powerful technique for exploring the embryonic development of the mammalian brain. In this protocol we describe a basic slice culture technique we have used for two sets of experiments: axon guidance transplant assays and bead culture assays.

Ex vivo Culture Assay Using Human Hair Follicles to Study Circadian Characteristics

Authors: Atsuhiro Nishida, Yoshiki Miyawaki, Koichi Node and Makoto Akashi, date: 06/05/2020, view: 1123, Q&A: 0

[Abstract] Ex vivo culture assays of biopsy specimens are advantageous for the experimental evaluation of human circadian characteristics. We developed a simple and non-invasive experimental evaluation method for monitoring the expression of circadian clock genes in an ex vivo culture assay using human hair follicles. This method imposes ...

FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells

Authors: Brandon T. Milliken, Robert P. Doyle, George G. Holz and Oleg G. Chepurny, date: 06/05/2020, view: 2062, Q&A: 0

[Abstract] Stimulation of G protein-coupled receptors (GPCR) by hormones and neurotransmitters elicits cellular responses, many of which result from alterations in the concentrations of cytosolic cAMP and Ca²⁺. Here, we describe a microplate reader fluorescence resonance energy transfer (FRET) assay that uses the genetically encoded biosensors ...

Measurement of Mitotic Spindle Angle and Mitotic Cell Distance in Fixed Tissue of Drosophila Larval Brains

Authors: Maribel Franco and Ana Carmena, date: 11/20/2019, view: 1691, Q&A: 0

[Abstract] The positioning and the cleavage plane orientation of mitotic cells in pseudostratified epithelia (PSE) must be tightly regulated since failures in any of these processes might have fatal consequences during development. Here we present a simple method to determine the spindle orientation as well as the positioning of neuroepithelial mitotic cells ...

Ex vivo Drosophila Wing Imaginal Disc Culture and Furin Inhibitor Assay

Authors: Alex Sohr, Lijuan Du and Sougata Roy, date: 08/20/2019, view: 2533, Q&A: 0

[Abstract] Furin is an evolutionarily conserved proprotein convertase (PC) family enzyme with a broad range of substrates that are essential for developmental, homeostatic, and disease pathways. Classical genetic approaches and in vitro biochemical or cell biological assays identified that precursor forms of most growth factor family proteins are ...

Lipid-exchange Rate Assay for Lipid Droplet Fusion in Live Cells

Authors: Jia Wang, Boon Tin Chua, Peng Li and Feng-Jung Chen, date: 07/20/2019, view: 2902, Q&A: 0

[Abstract] Lipid droplets (LDs) are central organelles in maintaining lipid homeostasis. Defective LD growth often results in the development of metabolic disorders. LD fusion and growth mediated by cell death–inducing DNA fragmentation factor alpha (DFFA)-like effector (CIDE) family proteins are crucial for various biological processes including unilocular ...

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