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Past Issues in 2015

Volume: 5 Issue: 11

Cell Biology

Dictyostelium Cultivation, Transfection, Microscopy and Fractionation Authors: Jennifer Hirst, Robert R Kay and David Traynor, date: 06/05/2015, view: 9010, Q&A: 0

The real time visualisation of fluorescently tagged proteins in live cells using ever more sophisticated microscopes has greatly increased our understanding of the dynamics of key proteins during fundamental physiological processes such as cell locomotion, chemotaxis, cell division and membrane trafficking. In addition the fractionation of cells ...

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FRAP Analysis of LET-23::GFP in the Vulval Epithelial Cells of Living Caenorhabditis elegans Larvae Authors: Michael Walser, Alex Hajnal and Juan M. Escobar-Restrepo, date: 06/05/2015, view: 5587, Q&A: 0

The Caenorhabditis elegans (C. elegans) vulva is a well-established system to study organ development as the molecular mechanisms that govern its formation are conserved in animals. Of special interest is the EGFR/RAS/MAPK signaling pathway that is required for fate acquisition and morphogenesis of the vulva. let-23 ...

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Immunology

Extraction and Identification of T Cell Stimulatory Self-lipid Antigens Authors: Marco Lepore, Sebastiano Sansano, Claudia de Lalla, Paolo Dellabona, Giulia Casorati, Gennaro De Libero and Lucia Mori, date: 06/05/2015, view: 5790, Q&A: 0

Autoreactive T cells restricted to CD1 molecules and specific for endogenous lipids are abundant in human blood (de Jong et al., 2010; de Lalla et al., 2011). A few self-lipid molecules recognized by diverse individual T cell clones and accumulated within APCs following stress signals or cell transformation have been identified ...

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Cytokine-Stimulated Phosphoflow of Whole Blood Using CyTOF Mass Cytometry Authors: Rosemary Fernandez and Holden Maecker, date: 06/05/2015, view: 8424, Q&A: 1

The ability to assess the function of a range of cytokine, antigen receptor, and Toll-like receptor (TLR) signaling pathways in a range of immune cells could provide a kind of fingerprint of the state of the human immune system. The mass cytometry or CyTOF, platform allows for the parallel application of about 40 labeled antibodies to a single ...

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Cytokine-stimulated Phosphoflow of PBMC Using CyTOF Mass Cytometry Authors: Rosemary Fernandez and Holden Maecker, date: 06/05/2015, view: 10784, Q&A: 1

Phosphorylation of tyrosine, serine, and threonine residues is critical for the control of protein activity involved in various cellular events. An assortment of kinases and phosphatases regulate intracellular protein phosphorylation in many different cell signaling pathways. These pathways include T and B cell signaling, regulating growth and ...

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Microbiology

Quantification of Total and 2-LTR (Long terminal repeat) HIV DNA, HIV RNA and Herpesvirus DNA in PBMCs Authors: Marta Massanella, Sara Gianella, Steven M. Lada, Douglas D. Richman and Matthew C. Strain, date: 06/05/2015, view: 7851, Q&A: 0

Almost all individuals infected with human immunodeficiency virus (HIV) are also infected with cytomegalovirus (CMV) and Epstein Barr virus (EBV). The aims of our studies have included characterizing and measuring the latent HIV reservoir and understanding the association between asymptomatic replication of CMV (and other herpesvirus, including ...

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Molecular Biology

DNA Damage Sensitivity Assays in Caenorhabditis elegans Authors: Hyun-Min Kim and Monica P. Colaiácovo, date: 06/05/2015, view: 7959, Q&A: 0

C. elegans has served as a genetically tractable multicellular model system to examine DNA damage-induced genotoxic stress which threatens genome integrity. Importantly, the high degree of conservation shared between worms and humans offers the advantage that findings about DNA damage-induced cell cycle arrest/checkpoint response and DNA ...

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Enzymatic Activity Assays for Base Excision Repair Enzymes in Cell Extracts from Vertebrate Cells Authors: Melike Çağlayan, Julie K. Horton and Samuel H. Wilson, date: 06/05/2015, view: 5218, Q&A: 0

We previously reported enzymatic activity assays for the base excision repair (BER) enzymes DNA polymerase β (pol β), aprataxin (APTX), and flap endonuclease 1 (FEN1) in cell extracts from Saccharomyces cerevisiae (Çağlayan and Wilson, 2014). Here, we describe a method to prepare cell extracts from vertebrate cells to investigate these ...

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Plant Science

Determination of the Developmental Origin of Seeds Containing Endosperm Using Flow Cytometric Analysis Authors: Christian Sailer, Anja Schmidt and Ueli Grossniklaus, date: 06/05/2015, view: 6112, Q&A: 0

Seeds derived from a diploid, sexual plant typically contain a 2n embryo (n+n) and 3n endosperm, a ratio characteristic for most flowering plants. However, this ratio is altered in apomictic species, which reproduce asexually through seeds (Koltunow and Grossniklaus, 2003). Apomixis is usually a facultative trait and encompasses several ...

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Magnaporthe oryzae Inoculation of Rice Seedlings by Spraying with a Spore Suspension Authors: Aya Akagi, Chang-Jie Jiang and Hiroshi Takatsuji, date: 06/05/2015, view: 8676, Q&A: 1

Fungal blast caused by Magnaporthe oryzae (M. oryzae) leads to a serious yield loss of rice. Appropriate assessment of disease occurrence is necessary to investigate the nature of the disease and plant strategies to resist the disease. We describe our assay method of disease severity of M. oryzae on intact rice leaves ...

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Simple Digital Photography for Assessing Biomass and Leaf Area Index in Cereals Authors: Jaume Casadesús and Dolors Villegas, date: 06/05/2015, view: 7639, Q&A: 0

These instructions refer to obtaining fast and low-labour estimates of ground cover, leaf area index and green biomass for a large number of plots, as those encountered in cereal breeding programs. The procedure includes obtaining the pictures in the field and processing them once downloaded to a computer.

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Measurement of Net High-affinity Urea Uptake in Maize Plants Authors: Laura Zanin, Nicola Tomasi and Roberto Pinton, date: 06/05/2015, view: 5574, Q&A: 0

Despite its extensive use as a nitrogen fertilizer, the role of urea as a directly accessible nitrogen source for crop plants is still poorly understood. So far, the physiological and molecular aspects of urea acquisition have been investigated only in a few plant species highlighting the importance of a urea transporter in roots, DUR3 (Kojima et ...

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Cell Wall Biomass Preparation and Fourier Transform Mid-infrared (FTIR) Spectroscopy to Study Cell Wall Composition Authors: Ricardo M. F. da Costa, Gordon G. Allison and Maurice Bosch, date: 06/05/2015, view: 5748, Q&A: 0

Plant cell wall biomass is an abundant and renewable organic resource. Of the polymers it encloses, cellulose and hemicellulose are regarded as a raw material for the production of fuels and other products (Klemm et al., 2005; Slavov et al., 2013). Nonetheless, current usage of lignocellulosic biomass is still below its full ...

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Quantitative Image Analysis of Membrane Microdomains Labelled by Fluorescently Tagged Proteins in Arabidopsis thaliana and Nicotiana benthamiana Authors: Iris K. Jarsch and Thomas Ott, date: 06/05/2015, view: 5692, Q&A: 0

We have recently characterized co-existing membrane microdomains that are labeled by different proteins in living plant cells (Jarsch et al., 2014). For this approach we first created a digital fingerprint for each of the twenty marker proteins using quantitative image analysis. Here we recorded parameters such as domain size, density and ...

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