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  • Volume 8, 2018
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Past Issues in 2018

Volume: 8 Issue: 4

Biochemistry

Characterization of Amyloid Fibril Networks by Atomic Force Microscopy Authors:  Mirren Charnley , Jay Gilbert, Owen G. Jones and Nicholas P. Reynolds, date: 02/20/2018, view: 801, Q&A: 0
Dense networks of amyloid nanofibrils fabricated from common globular proteins adsorbed to solid supports can improve cell adhesion, spreading and differentiation compared to traditional flat, stiff 2D cell culture substrates like Tissue Culture Polystyrene (TCPS). This is due to the fibrous, nanotopographic nature of the amyloid fibril networks ...
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Cell Biology

Sebinger Culture: A System Optimized for Morphological Maturation and Imaging of Cultured Mouse Metanephric Primordia Authors:  Mona Elhendawi and Jamie A. Davies, date: 02/20/2018, view: 633, Q&A: 0
Here, we present a detailed protocol on setting up embryonic renal organ cultures using a culture method that we have optimised for anatomical maturation and imaging. Our culture method places kidney rudiments on glass in a thin film of medium, which results in very flat cultures with all tubules in the same image plane. For reasons not yet ...
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Microbiology

A Small RNA Isolation and Sequencing Protocol and Its Application to Assay CRISPR RNA Biogenesis in Bacteria Authors:  Sukrit Silas, Nimit Jain, Michael Stadler, Becky Xu Hua Fu, Antonio Sánchez-Amat, Andrew Z. Fire and Joshua Arribere, date: 02/20/2018, view: 994, Q&A: 0
Next generation high-throughput sequencing has enabled sensitive and unambiguous analysis of RNA populations in cells. Here, we describe a method for isolation and strand-specific sequencing of small RNA pools from bacteria that can be multiplexed to accommodate multiple biological samples in a single experiment. Small RNAs are isolated by ...
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Conditional Knockdown of Proteins Using Auxin-inducible Degron (AID) Fusions in Toxoplasma gondii Authors:  Kevin M. Brown, Shaojun Long and L. David Sibley, date: 02/20/2018, view: 1528, Q&A: 0
Toxoplasma gondii is a member of the deadly phylum of protozoan parasites called Apicomplexa. As a model apicomplexan, there is a great wealth of information regarding T. gondii’s 8,000+ protein coding genes including sequence variation, expression, and relative contribution to parasite fitness. However, new tools are needed to ...
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Flow Cytometric Quantification of Fatty Acid Uptake by Mycobacterium tuberculosis in Macrophages Authors:  Evgeniya V. Nazarova, Maria Podinovskaia, David G. Russell and Brian C. VanderVen, date: 02/20/2018, view: 806, Q&A: 0
Mycobacterium tuberculosis (Mtb) has evolved to assimilate fatty acids from its host. However, until recently, there was no reliable way to quantify fatty acid uptake by the bacteria during host cell infection. Here we describe a new method to quantify fatty acid uptake by intracellular bacilli. We infect macrophages with Mtb ...
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Molecular Biology

An Improved Method for Measuring Chromatin-binding Dynamics Using Time-dependent Formaldehyde Crosslinking Authors:  Elizabeth A. Hoffman, Hussain Zaidi, Savera J. Shetty, Stefan Bekiranov and David T. Auble, date: 02/20/2018, view: 909, Q&A: 0
Formaldehyde crosslinking is widely used in combination with chromatin immunoprecipitation (ChIP) to measure the locations along DNA and relative levels of transcription factor (TF)-DNA interactions in vivo. However, the measurements that are typically made do not provide unambiguous information about the dynamic properties of these ...
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Neuroscience

Registration and Alignment Between in vivo Functional and Cytoarchitectonic Maps of Mouse Visual Cortex Authors:  Jun Zhuang, Quanxin Wang, Marc Takeno and Jack Waters, date: 02/20/2018, view: 485, Q&A: 0
This protocol describes a method for registration of in vivo cortical retinotopic map with cytochrome c oxidase (CO) labeled architectonic maps of the same mouse brain through the alignment of vascular fiducials. By recording surface blood vessel pattern and sequential alignment at each step, this method overcomes the challenge imposed by ...
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Assaying Mechanonociceptive Behavior in Drosophila Larvae Authors:  Nina Hoyer, Meike Petersen, Federico Tenedini and Peter Soba, date: 02/20/2018, view: 730, Q&A: 0
Drosophila melanogaster larvae have been extensively used as a model to study the molecular and cellular basis of nociception. The larval nociceptors, class IV dendritic arborization (C4da) neurons, line the body wall of the animal and respond to various stimuli including noxious heat and touch. Activation of C4da neurons results in a ...
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Assaying Thermo-nociceptive Behavior in Drosophila Larvae Authors:  Meike Petersen, Federico Tenedini, Nina Hoyer, Fritz Kutschera and Peter Soba, date: 02/20/2018, view: 668, Q&A: 0
Thermo-nociception, the detection and behavioral response to noxious temperatures, is a highly conserved action to avoid injury and ensure survival. Basic molecular mechanisms of thermal responses have been elucidated in several model organisms and are of clinical relevance as thermal hypersensitivity (thermos-allodynia) is common in neuropathic ...
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Plate Assay to Determine Caenorhabditis elegans Response to Water Soluble and Volatile Chemicals Authors:  Takashi Murayama and Ichiro N. Maruyama, date: 02/20/2018, view: 699, Q&A: 0
The nematode Caenorhabditis elegans is widely used for behavioral studies ranging from simple chemosensation to associative learning and memory. It is vital for such studies to determine optimal concentrations of attractive and aversive chemicals that C. elegans can sense. Here we describe a resource localization assay in which a ...
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Plant Science

Transient Gene Expression for the Characteristic Signal Sequences and the Estimation of the Localization of Target Protein in Plant Cell Authors:  Mikhail Berestovoy, Alexander Tyurin, Ksenia Kabardaeva, Yuriy Sidorchuk, Artem Fomenkov, Alexander Nosov and Irina Goldenkova-Pavlova, date: 02/20/2018, view: 749, Q&A: 0
We have proposed and tested a method for characterization of the signal sequences and determinations of target protein localization in a plant cell. This method, called the AgI-PrI, implies extraction of protoplasts from plant tissues after agroinfiltration. The suggested approach combines the advantages of two widely used methods for transient ...
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Measurement of Arabidopsis thaliana Plant Traits Using the PHENOPSIS Phenotyping Platform Authors:  Wojciech Rymaszewski, Myriam Dauzat, Alexis Bédiée, Gaëlle Rolland, Nathalie Luchaire, Christine Granier, Jacek Hennig and Denis Vile, date: 02/20/2018, view: 682, Q&A: 0
High-throughput phenotyping of plant traits is a powerful tool to further our understanding of plant growth and its underlying physiological, molecular, and genetic determinisms. This protocol describes the methodology of a standard phenotyping experiment in PHENOPSIS automated platform, which was engineered in INRA-LEPSE ( ...
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Stem Cell

Preparation of Amyloid Fibril Networks Authors:  Mirren Charnley , Jay Gilbert, Owen G. Jones and Nicholas P. Reynolds, date: 02/20/2018, view: 623, Q&A: 0
Networks of amyloid nanofibrils fabricated from common globular proteins such as lysozyme and β-lactoglobulin have material properties that mimic the extracellular microenvironment of many cell types. Cells cultured on such amyloid fibril networks show improved attachment, spreading and in the case of mesenchymal stem cells improved ...
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Isolation and Establishment of Mesenchymal Stem Cells from Wharton’s Jelly of Human Umbilical Cord Authors:  Umesh Goyal, Chitra Jaiswal and Malancha Ta, date: 02/20/2018, view: 1171, Q&A: 3
Mesenchymal stem cells (MSCs) are currently considered as ‘medicinal signaling cells’ and a promising resource in regard to cell-based regenerative therapy. Umbilical cord is a human term perinatal tissue which is easily attainable, and a promising source of stem cells with no associated ethical concerns. MSCs have been isolated from different ...
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Systems Biology

Adapting the Smart-seq2 Protocol for Robust Single Worm RNA-seq Authors:  Lorrayne Serra, Dennis Z. Chang, Marissa Macchietto, Katherine Williams, Rabi Murad, Dihong Lu, Adler R. Dillman and Ali Mortazavi, date: 02/20/2018, view: 1236, Q&A: 0
Most nematodes are small worms that lack enough RNA for regular RNA-seq protocols without pooling hundred to thousand of individuals. We have adapted the Smart-seq2 protocol in order to sequence the transcriptome of an individual worm. While developed for individual Steinernema carpocapsae and Caenorhabditis elegans larvae as ...
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