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Past Issues in 2014

Volume: 4 Issue: 20

Immunology

Measurement of TACE Activity in Extracts from Cultured Cells Authors: Xiuling Xu, Mabel T. Padilla and Yong Lin, date: 10/20/2014, view: 4932, Q&A: 0

In cigarette smoke–induced and inflammation-associated lung cancer development, cigarette smoke extract (CSE) activates tumor necrosis factor-alpha (TNF-α) secretion from macrophages. TNF-α converting enzyme (TACE), also known as α-Secretase or ADAM17 (A Disintegrin and Metalloprotease), is a member of the ADAM family of metalloproteases. TACE ...

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Microbiology

Extraction and Purification of Mycobacterial Mycolic Acids Authors: Christian M. Dupont and Laurent Kremer, date: 10/20/2014, view: 7285, Q&A: 0

Mycolic acids are major long-chain fatty acids, containing up to 80-90 carbon atoms that represent essential components of the mycobacterial cell wall (Pawelczyk and Kremer, 2014). Each mycobacterial species possesses a specific mycolic acid profile characterized by various chemical modifications that decorate the lipid. Mycolic acids play a ...

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In vitro Assay for Cytidine Deaminase Activity of APOBEC3 Protein Authors: Smita Nair and Alan Rein, date: 10/20/2014, view: 6905, Q&A: 0

Cytidine deaminases are enzymes that catalyze the removal of an amino group from cytidine, forming uridine. APOBEC3 (ApolipoproteinB mRNA editing enzyme, catalytic polypeptide like) proteins are cytidine deaminases that deaminate cytidines in polynucleotides (RNA/DNA), resulting in editing of their target substrates. Mammalian APOBEC3 proteins are ...

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Design of a Transcription-based Secretion Activity Reporter (TSAR) for the Type III Secretion Apparatus of Shigella flexneri and Uses Thereof Authors: François Xavier Campbell-Valois, Pamela Schnupf and Philippe J. Sansonetti, date: 10/20/2014, view: 5558, Q&A: 1

Many gram-negative bacterial pathogens, including Shigella flexneri, are able to translocate bacterial proteins, dubbed effectors, across the host cell plasma membrane into the host cell cytosol using a syringe-like structure, the type three secretion apparatus (T3SA). While some bacteria use their T3SA to modulate their phagosomal ...

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Detection of the Secreted and Cytoplasmic Fractions of IpaB, IpaC and IpaD by Lysozyme Permeabilization Authors: François Xavier Campbell-Valois, Pamela Schnupf and Philippe J. Sansonetti, date: 10/20/2014, view: 5035, Q&A: 0

Gram negative bacterial pathogens, such as Shigella flexneri, which possess a Type Three Secretion System (T3SS), are able to transfer bacterial proteins, dubbed translocators and effectors, from their cytoplasm into the cytoplasm of their host cells using a syringe like needle complex. For Shigella, it has been shown that during ...

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Measuring UV-induced Mutagenesis at the CAN1 Locus in Saccharomyces cerevisiae Authors: Ildiko Unk and Andreea Daraba, date: 10/20/2014, view: 5928, Q&A: 1

There are several methods to measure the capacity of yeast cell to respond to environmental impacts on their genome by mutating it. One frequently used method involves the detection of forward mutations in the CAN1 gene. The CAN1 gene encodes for an arginine permease that is responsible for the uptake of arginine and it can also ...

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Synchronization of Saccharomyces cerevisiae Cells in G1 Phase of the Cell Cycle Authors: Ildiko Unk and Andreea Daraba, date: 10/20/2014, view: 8804, Q&A: 0

The baker’s yeast, Saccharomyces cerevisiae is a widely used model organism in molecular biology because of the high homology it shares with human cells in many basic cellular processes such as DNA replication, repair, recombination, transcription, and because of the ease its genome can be manipulated. Other advantages of working with ...

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Severe Fever with Thrombocytopenia Syndrome Virus Infection of Cell Cultures Authors: Yinyan Sun and Wenhui Li, date: 10/20/2014, view: 4560, Q&A: 0

Severe fever with thrombocytopenia syndrome virus (SFTSV) can infect multiple cells, such as Vero, RD, BHK21, HUVEC and 293T. Vero cells are highly susceptible, so they were used to isolate virus from sera of patients, as well as for producing virus in the laboratory. Other cells can be used for additional functional analysis.

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Molecular Biology

Expression and Purification of the Eukaryotic MBP-MOS1 Transposase from Sf21 Insect Cells Authors: Jérôme Jaillet, Audrey Dussaussois-Montagne, Sylvaine Renault and Corinne Augé-Gouillou, date: 10/20/2014, view: 6031, Q&A: 0

Here, we present the full-length protocol for purifying the recombinant MOS1 transposase from insect cells used in our recent publication (Pflieger et al., 2014), which involved a N-terminal MBP-tag and maltose-affinity chromatography.

Due to their overall basic properties, transposases are often difficult to purify, ...

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Neuroscience

Olfactory Habituation in Fasted Mice Authors: Tifany Desprez, Giovanni Marsicano and Edgar Soria-Gomez, date: 10/20/2014, view: 4783, Q&A: 0

Sensory perception is tightly modulated by the individual’s internal states. In particular, it has been shown that olfactory processes are constantly influenced by metabolic signals reflecting the energy status of the body. Thus, it is important to implement novel approaches to evaluate the impact of body energy changes on olfactory performance. ...

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Plant Science

Plastic Embedding of Arabidopsis Stem Sections Authors: Florian Chevalier, Soledad Montalbán Iglesias, Óscar Javier Sánchez, Lluís Montoliu and Pilar Cubas, date: 10/20/2014, view: 7180, Q&A: 0

The inflorescence stem of the flowering plant Arabidopsis thaliana (thale cress) is an excellent model system to investigate plant vascular tissue patterning and development. Plant vasculature is a complex conducting tissue arranged in strands called vascular bundles, formed by xylem (tissue that carries water) and phloem (tissue that ...

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Seed Storage Reserve Analysis Authors: Harrie van Erp, Guillaume Menard and Peter J. Eastmond, date: 10/20/2014, view: 4565, Q&A: 0

One of the major goals of plant research is to improve crop yield, by for instance increasing seed oil or protein content. Besides this, extensive research is done to change seed fatty acid (FA) composition in order to make vegetable oils more suitable for specific purposes. To determine the effect of genetic changes on seed FA composition, oil, ...

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Fluorescence Recovery after Photobleaching (FRAP) Assay to Measure the Dynamics of Fluorescence Tagged Proteins in Endoplasmic Reticulum Membranes of Plant Cells Authors: José Antonio Navarro, Marta Serra-Soriano and Vicente Pallás, date: 10/20/2014, view: 8243, Q&A: 0

In this protocol, we used fluorescence recovery after photobleaching (FRAP) to measure the influence that some mutations and drug treatment have on mobility of a green fluorescent protein (GFP)-fused viral transmembrane protein into endoplasmic reticulum membranes (Serra-Soriano et al., 2014). The proteins of interest were transiently ...

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A Protocol to Measure the Extent of Cell-to-cell Movement of RNA Viruses in Planta Authors: José Antonio Navarro, Marta Serra-Soriano and Vicente Pallás, date: 10/20/2014, view: 4002, Q&A: 0

Here, we present a simple and rapid protocol to measure the extent of cell-to-cell movement of RNA viruses in planta. To do that, the green fluorescent protein (GFP) gene was incorporated into the genome of Melon necrotic spot virus (MNSV) as a coat protein (CP) fusion protein using the Thosea asigna virus 2A catalytic ...

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