• Volume 11, 2021
  • Volume 10, 2020
  • Volume 9, 2019
  • Volume 8, 2018
  • Volume 7, 2017
  • Volume 6, 2016
  • Volume 5, 2015
  • Volume 4, 2014
  • Volume 3, 2013
  • Volume 2, 2012
  • Volume 1, 2011
Image Credit: BioProtoc.4105

Current Issue in 2021

Volume: 11, Issue: 15

Biochemistry

Efficient and Rapid Analysis of Polysomes and Ribosomal Subunits in Cells and Tissues Using Ribo Mega-SEC Authors:  Harunori Yoshikawa, Ramasubramanian Sundaramoorthy, Daniel Mariyappa, Hao Jiang and Angus I. Lamond, date: 08/05/2021, view: 20, Q&A: 0

Polysome profile analysis is a popular method for separating polysomes and ribosomal subunits and is typically achieved using a sucrose density gradient (SDG). This has remained the gold standard method since ribosomes were first discovered; however, this method is time-consuming and requires multiple steps from making the gradient and long

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Purification of Mitochondrial Ribosomes with the Translocase Oxa1L from HEK Cells Authors:  Hanting Yang and Nirupa Desai, date: 08/05/2021, view: 30, Q&A: 0

Mitochondrial ribosomes (mitoribosomes) perform protein synthesis inside mitochondria, the organelles responsible for energy conversion and adenosine triphosphate (ATP) production in eukaryotic cells. To investigate their functions and structures, large-scale purification of intact mitoribosomes from mitochondria-rich animal tissues or HEK cells

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Purification of Recombinant Wild Type and Mutant Ryanodine Receptors Expressed in HEK293 Cells Authors:  Yifan Hu, Kavita A. Iyer, Ashok R. Nayak, Nagomi Kurebayashi, Takashi Murayama and Montserrat Samsó, date: 08/05/2021, view: 22, Q&A: 0

High quantities of purified ryanodine receptor (RyR), a large (2.26 MDa) intracellular homotetrameric membrane protein, can be obtained from heterologous expression in HEK293 cells and used for structure determination by cryo-EM. The advantage of using recombinant protein is that the variability due to post-translational modifications can be

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Biophysics

Purification and Cryo-electron Microscopy Analysis of Plant Mitochondrial Ribosomes Authors:  Florent Waltz, Philippe Giegé and Yaser Hashem, date: 08/05/2021, view: 17, Q&A: 0

Plants make up by far the largest part of biomass on Earth. They are the primary source of food and the basis of most drugs used for medicinal purposes. Similarly to all eukaryotes, plant cells also use mitochondria for energy production. Among mitochondrial gene expression processes, translation is the least understood; although, recent advances

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Cell Biology

CRISPR-mediated Labeling of Cells in Chick Embryos Based on Selectively Expressed Genes Authors:  Masahito Yamagata and Joshua R. Sanes, date: 08/05/2021, view: 62, Q&A: 0

The abilities to mark and manipulate specific cell types are essential for an increasing number of functional, structural, molecular, and developmental analyses in model organisms. In a few species, this can be accomplished by germline transgenesis; in other species, other methods are needed to selectively label somatic cells based on the genes

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Cell-attached and Whole-cell Patch-clamp Recordings of Dopamine Neurons in the Substantia Nigra Pars Compacta of Mouse Brain Slices Authors:  Stefano Cattaneo, Maria Regoni, Jenny Sassone and Stefano Taverna, date: 08/05/2021, view: 17, Q&A: 0

The Substantia Nigra pars compacta (SNc) is a midbrain dopaminergic nucleus that plays a key role in modulating motor and cognitive functions. It is crucially involved in several disorders, particularly Parkinson’s disease, which is characterized by a progressive loss of SNc dopaminergic cells. Electrophysiological studies on SNc neurons are

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Developmental Biology

Isolation and in vitro Culture of Mouse Oocytes Authors:  Jessica Greaney, Goutham Narayanan Subramanian, Yunan Ye and Hayden Homer, date: 08/05/2021, view: 23, Q&A: 0

Females are endowed at birth with a fixed reserve of oocytes, which declines both in quantity and quality with advancing age. Understanding the molecular mechanisms regulating oocyte quality is crucial for improving the chances of pregnancy success in fertility clinics. In vitro culture systems enable researchers to analyse important molecular and

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A Genetically Engineered Mouse Model of Venous Anomaly and Retinal Angioma-like Vascular Malformation Authors:  Xudong Cao, Beibei Xu, Xiao Li, Taotao Li and Yulong He, date: 08/05/2021, view: 16, Q&A: 0

Characterization of key regulators in vein development will advance our understanding of mechanisms underlying venous anomalies and provide therapeutic targets for the treatment of vascular malformations. Here, we provide a detailed protocol for the generation of genetically engineered mouse models targeting the Tek gene for the analysis of vein

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Medicine

A Swimming-based Assay to Determine the Exercise Capacity of Adult Zebrafish Cardiomyopathy Models Authors:  Xiao Ma and Xiaolei Xu, date: 08/05/2021, view: 18, Q&A: 0

Exercise capacity, measured by treadmill in humans and other mammals, is an important diagnostic and prognostic index for patients with cardiomyopathy and heart failure. The adult zebrafish is increasingly used as a vertebrate model to study human cardiomyopathy due to its conserved cardiovascular physiology, convenience for genetic manipulation,

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Microbiology

Ion Transport Activity Assay for Microbial Rhodopsin Expressed in Escherichia coli Cells Authors:  Masae Konno, Keiichi Inoue and Hideki Kandori, date: 08/05/2021, view: 28, Q&A: 0

Microbial rhodopsins have diverse functions, including roles as light-driven ion pumps, light-gated ion channels, photosensors, and light-regulated enzymes. As the number of rhodopsin-like genes identified has increased in recent years, so has the requirement for rapid identification of their functions. The patch-clamp method is often used to

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Molecular Biology

Isolation of Nuclei from Mouse Dorsal Root Ganglia for Single-nucleus Genomics Authors:  Lite Yang, Ivan Tochitsky, Clifford J. Woolf and William Renthal, date: 08/05/2021, view: 24, Q&A: 0

Primary somatosensory neurons, whose cell bodies reside in the dorsal root ganglion (DRG) and trigeminal ganglion, are specialized to transmit sensory information from the periphery to the central nervous system. Our molecular understanding of peripheral sensory neurons has been limited by both their heterogeneity and low abundance compared with

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Fe-NTA Microcolumn Purification of Phosphopeptides from Immunoprecipitation (IP) Eluates for Mass Spectrometry Analysis Authors:  Ethan J. Sanford and Marcus B. Smolka, date: 08/05/2021, view: 14, Q&A: 0

Protein phosphorylation is a nearly universal signaling mechanism. To date, a number of proteomics tools have been developed to analyze phosphorylation. Phosphoproteome-wide analyses using whole cell extracts suffer from incomplete coverage, often missing phosphorylation events from low-abundance proteins. In order to increase coverage of

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Neuroscience

Optical Clearing and 3D Analysis Optimized for Mouse and Human Pancreata Authors:  Alexandra Alvarsson, Maria Jimenez-Gonzalez, Rosemary Li, Carolina Rosselot, Nikolaos Tzavaras, Zhuhao Wu and Sarah A. Stanley, date: 08/05/2021, view: 44, Q&A: 0

The pancreas is a heavily innervated organ, but pancreatic innervation can be challenging to comprehensively assess using conventional histological methods. However, recent advances in whole-mount tissue clearing and 3D rendering techniques have allowed detailed reconstructions of pancreatic innervation. Optical clearing is used to enhance tissue

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A Simple Spatial-independent Associative and Reversal Learning Task in Mice Authors:  Fulvio Magara, Benjamin Boury-Jamot and Hanna Hörnberg, date: 08/05/2021, view: 17, Q&A: 0

The ability to adapt one's behavior in response to changing circumstances, or cognitive flexibility, is often altered in neuropsychiatric and neurodevelopmental conditions. In rodents, cognitive flexibility is frequently assessed using associative learning paradigms with a reversal component. The majority of existing protocols rely on

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Plant Science

A Fast and Easy Method to Study Ralstonia solanacearum Virulence upon Transient Gene Expression or Gene Silencing in Nicotiana benthamiana Leaves Authors:  Wenjia Yu and Alberto P. Macho, date: 08/05/2021, view: 20, Q&A: 0

Ralstonia solanacearum is a devastating soil-borne bacterial pathogen that causes disease in multiple host plants worldwide. Typical assays to measure virulence of R. solanacearum in laboratory conditions rely on soil-drenching inoculation followed by observation and scoring of disease symptoms. Here, we describe a novel inoculation protocol to

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Stem Cell

Mouse Periosteal Cell Culture, in vitro Differentiation, and in vivo Transplantation in Tibial Fractures Authors:  Simon Perrin, Anais Julien, Oriane Duchamp de Lageneste, Rana Abou-Khalil and Céline Colnot, date: 08/05/2021, view: 46, Q&A: 0

The periosteum covering the outer surface of bone contains skeletal stem/progenitor cells that can efficiently form cartilage and bone during bone repair. Several methods have been described to isolate periosteal cells based on bone scraping and/or enzymatic digestion. Here, we describe an explant culture method to isolate periosteum-derived

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