Volume: 8 Issue: 19 —BIO-PROTOCOL

Biochemistry

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: 3365, Q&A: 0

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, ...

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Expression and Ni-NTA-Agarose Purification of Recombinant Hepatitis C Virus E2 Ectodomain Produced in a Baculovirus Expression System Authors: Julián Gómez-Gutiérrez, Mar Rodríguez-Rodríguez, Francisco Gavilanes and Belén Yélamos, date: 10/05/2018, view: 2541, Q&A: 0

In this protocol, we describe the production and purification of the ectodomain of the E2₆₆₁ envelope protein (amino acids 384-661) of the Hepatitis C virus, which plays a fundamental role in the entry of the virus into the host cell. This protein has been expressed in both prokaryotic and eukaryotic systems but in small quantities or ...

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A Method for SUMO Modification of Proteins in vitro Authors: Christine C. Lee, Bing Li, Hongtao Yu and Michael J. Matunis, date: 10/05/2018, view: 3240, Q&A: 0

The Small Ubiquitin-related Modifier (SUMO) is a protein that is post-translationally added to and reversibly removed from other proteins in eukaryotic cells. SUMO and enzymes of the SUMO pathway are well conserved from yeast to humans and SUMO modification regulates a variety of essential cellular processes including transcription, chromatin ...

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Filter Retardation Assay for Detecting and Quantifying Polyglutamine Aggregates Using Caenorhabditis elegans Lysates Authors: Olga Sin, Alejandro Mata-Cabana, Renée I. Seinstra and Ellen A. A. Nollen, date: 10/05/2018, view: 4163, Q&A: 0

Protein aggregation is a hallmark of several neurodegenerative diseases and is associated with impaired protein homeostasis. This imbalance is caused by the loss of the protein’s native conformation, which ultimately results in its aggregation or abnormal localization within the cell. Using a C. elegans model of polyglutamine diseases, we ...

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Cell Biology

Maintenance of Schmidtea mediterranea in the Laboratory Authors: Nídia de Sousa and Teresa Adell, date: 10/05/2018, view: 3204, Q&A: 0

In the last years, planarians have emerged as a unique model animal for studying regeneration and stem cells biology. Although their remarkable regenerative abilities are known for a long time, only recently the molecular tools to understand the biology of planarian stem cells and the fundamentals of their regenerative process have been ...

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Developmental Biology

Detection of Cell Death in Planarians Authors: Nídia de Sousa and Teresa Adell, date: 10/05/2018, view: 3271, Q&A: 0

Planarians are freshwater flatworms, well known for their ability to regenerate a complete organism from any piece of their body. Furthermore, planarians are constantly growing and degrowing throughout their lives, maintaining a functional and proportioned body. These properties rely on the presence of a population of adult stem cells and on the ...

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Immunology

Isolation and Culture of Mouse Lung ILC2s Authors: Pauline Schmitt, Anais Duval, Emilie Mirey, Jean-Philippe Girard and Corinne Cayrol, date: 10/05/2018, view: 4220, Q&A: 0

Group 2 Innate Lymphoid Cells (ILC2) play an important role in immune responses at barrier surfaces, notably in the lung during airway allergic inflammation or asthma. Several studies have described methods to isolate ILC2s from wild-type naive mice, most of them using cell sorting to obtain a pure population. Here, we describe in detail, a ...

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Microbiology

CRISPR/Cas9-mediated ssDNA Recombineering in Corynebacterium glutamicum Authors: Jiao Liu, Yu Wang, Ping Zheng and Jibin Sun, date: 10/05/2018, view: 3328, Q&A: 0

Corynebacterium glutamicum is a versatile workhorse for industrial bioproduction of many kinds of chemicals and fuels, notably amino acids. Development of advanced genetic engineering tools is urgently demanded for systems metabolic engineering of C. glutamicum. Recently unveiled clustered regularly interspaced short palindromic ...

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Molecular Biology

Fluorescence Titrations to Determine the Binding Affinity of Cyclic Nucleotides to SthK Ion Channels Authors: Philipp A.M. Schmidpeter and Crina M. Nimigean, date: 10/05/2018, view: 4480, Q&A: 0

The cyclic-nucleotide modulated ion channel family includes cyclic nucleotide-gated (CNG) and hyperpolarization-activated and cyclic nucleotide-modulated (HCN) channels, which play essential roles in visual and olfactory signaling and the heart pacemaking activity. Functionally, these channels have been extensively characterized by ...

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Neuroscience

Measurement of Dopamine Using Fast Scan Cyclic Voltammetry in Rodent Brain Slices Authors: Madelyn I. Mauterer, Paige M. Estave, Katherine M. Holleran and Sara R. Jones, date: 10/05/2018, view: 5051, Q&A: 0

Fast scan cyclic voltammetry (FSCV) is an electrochemical technique that allows sub-second detection of oxidizable chemical species, including monoamine neurotransmitters such as dopamine, norepinephrine, and serotonin. This technique has been used to record the physiological dynamics of these neurotransmitters in brain tissue, including their ...

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Isolation of Chromatin-bound Proteins from Subcellular Fractions for Biochemical Analysis Author: Sébastien Gillotin, date: 10/05/2018, view: 9136, Q&A: 3

Shuttling of proteins between different cellular compartments controls their proteostasis and can contribute in some cases to regulate their activity. Biochemical analysis of chromatin-bound proteins, such as transcription factors, is often difficult because of their low yield and due to the interference from nucleic acids. This protocol describes ...

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Drosophila Endurance Training and Assessment of Its Effects on Systemic Adaptations Authors: Deena Damschroder , Tyler Cobb, Alyson Sujkowski and Robert Wessells, date: 10/05/2018, view: 3325, Q&A: 0

Exercise induces beneficial systemic adaptations that reduce the incidence of age-related diseases. However, the molecular pathways that elicit these adaptations are not well understood. Understanding the molecular mechanisms that underlie the exercise response can lead to widely beneficial therapies. Large populations, relatively short lifespan, ...

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Plant Science

Real-time PCR Analysis of PAMP-induced Marker Gene Expression in Nicotiana benthamiana Authors: Fan Liu, Yuanpeng Xu, Yan Wang and Yuanchao Wang, date: 10/05/2018, view: 3612, Q&A: 0

Perception of pathogen-associated molecular patterns (PAMPs) often triggers various innate immune responses in plants. The transcriptional changes of defense-related genes are often used as a marker to assay PAMP-triggered plant immune response. Here we described a protocol to monitor the relative expression level of marker genes in Nicotiana ...

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An Image Analysis Pipeline to Quantify Emerging Cracks in Materials or Adhesion Defects in Living Tissues Authors: Stéphane Verger, Guillaume Cerutti and Olivier Hamant, date: 10/05/2018, view: 2931, Q&A: 0

Microcracks in materials reflect their mechanical properties. The quantification of the number or orientation of such cracks is thus essential in many fields, including engineering and geology. In biology, cracks in soft tissues can reflect adhesion defects, and the analysis of their pattern can help to deduce the magnitude and orientation of ...

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Past Issues in 2018

Volume: 8 Issue: 19

Biochemistry

Cell Biology

Developmental Biology

Immunology

Microbiology

Molecular Biology

Neuroscience

Plant Science