Featured protocol,
Authors: Imre Gáspár 
Imre GáspárAffiliation: European Molecular Biology Laboratory (EMBL), Developmental Biology Unit, Heidelberg, Meyerhofstrasse 1, D-69117, Germany
For correspondence: imre.gaspar@embl.deBio-protocol author page: a4784 Anne EphrussiAffiliation: European Molecular Biology Laboratory (EMBL), Developmental Biology Unit, Heidelberg, Meyerhofstrasse 1, D-69117, Germany
For correspondence: anne.ephrussi@embl.deBio-protocol author page: a4785
DOI: https://doi.org/10.21769/BioProtoc.2380.
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| Brief version appeared in EMBO J, Feb 2017 |
Understanding the dynamic behavior and the continuously changing composition of macromolecular complexes, subcellular structures and organelles is one of areas of active research in both cell and developmental biology, as these changes directly relate to function and subsequently to the development and homeostasis of the organism. Here, we demonstrate the use of the developing
Drosophila oocyte to study dynamics of messenger ribonucleoprotein complexes (mRNPs) with high spatiotemporal resolution. The combination of
Drosophila genetics with total internal reflection (TIRF) microscopy, image processing and data analysis gives insight into mRNP motility and composition dynamics with unprecedented precision.
Featured protocol,
Authors: Collin Yvès Ewald Collin Yvès EwaldAffiliation 1: Department of Health Sciences and Technology, Eidgenössische Technische Hochschule (ETH) Zürich, Schwerzenbach-Zürich, Switzerland
Affiliation 2: Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
Affiliation 3: Harvard Stem Cell Institute, Harvard University, Boston, Massachusetts, USA
Affiliation 4: Joslin Diabetes Center, Research Division, Boston, Massachusetts, USA
For correspondence: collin-ewald@ethz.chBio-protocol author page: a4749 John M. HourihanAffiliation 1: Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
Affiliation 2: Harvard Stem Cell Institute, Harvard University, Boston, Massachusetts, USA
Affiliation 3: Joslin Diabetes Center, Research Division, Boston, Massachusetts, USA
Bio-protocol author page: a4750 T. Keith BlackwellAffiliation 1: Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
Affiliation 2: Harvard Stem Cell Institute, Harvard University, Boston, Massachusetts, USA
Affiliation 3: Joslin Diabetes Center, Research Division, Boston, Massachusetts, USA
Bio-protocol author page: a4751
DOI: https://doi.org/10.21769/BioProtoc.2365.
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| Brief version appeared in Elife, Jan 2017 |
Cells and organisms face constant exposure to reactive oxygen species (ROS), either from the environment or as a by-product from internal metabolic processes. To prevent cellular damage from ROS, cells have evolved detoxification mechanisms. The activation of these detoxification mechanisms and their downstream responses represent an overlapping defense response that can be tailored to different sources of ROS to adequately adapt and protect cells. In this protocol, we describe how to measure the sensitivity to oxidative stress from two different sources, arsenite and tBHP, using the nematode
C. elegans.
Featured protocol,
Authors: Jun-yi Zhu*Jun-yi ZhuAffiliation: Center for Cancer and Immunology Research, Children’s National Medical Center, 111 Michigan Ave. NW, Washington, DC, USA
Bio-protocol author page: a4710 Yulong FuAffiliation: Center for Cancer and Immunology Research, Children’s National Medical Center, 111 Michigan Ave. NW, Washington, DC, USA
Bio-protocol author page: a4711 Adam RichmanAffiliation: Center for Cancer and Immunology Research, Children’s National Medical Center, 111 Michigan Ave. NW, Washington, DC, USA
Bio-protocol author page: a4712 Zhe HanAffiliation 1: Center for Cancer and Immunology Research, Children’s National Medical Center, 111 Michigan Ave. NW, Washington, DC, USA
Affiliation 2: Department of Pediatrics, The George Washington University School of Medicine and Health Sciences, Washington, DC, USA
For correspondence: zhan@childrensnational.orgBio-protocol author page: a4713
DOI: https://doi.org/10.21769/BioProtoc.2350.
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| Brief version appeared in Elife, Jan 2017 |
Genomic sequencing efforts can implicate large numbers of genes and
de novo mutations as potential disease risk factors. A high throughput
in vivo model system to validate candidate gene association with pathology is therefore useful. We present such a system employing
Drosophila to validate candidate congenital heart disease (CHD) genes. The protocols exploit comprehensive libraries of UAS-GeneX-RNAi fly strains that when crossed into a 4XHand-Gal4 genetic background afford highly efficient cardiac-specific knockdown of endogenous fly orthologs of human genes. A panel of quantitative assays evaluates phenotypic severity across multiple cardiac parameters. These include developmental lethality, larva and adult heart morphology, and adult longevity. These protocols were recently used to evaluate more than 100 candidate CHD genes implicated by patient whole-exome sequencing (Zhu
et al., 2017).