Development and optimization of a miniaturized western blot-based screening platform to identify regulators of post-translational modifications

Post-translational modifications (PTMs) are fundamental traits of protein functionality and their study has been addressed using several approaches over the past years. However, screening methods developed to detect regulators of PTMs imply many challenges and are usually based on expensive techniques. Herein, we described the development and optimization of a western blot-based platform for identification of regulators of a specific PTM—mono-ubiquitylation of proliferating cell nuclear antigen (PCNA). This cell-based method does not require specific equipment, apart from the basic western blot (WB) devices and minor accessories, which are accessible for most research labs. The modifications introduced to the classical WB protocol allow the performance of PTM analysis from a single well of a 96-well plate with minimal sample manipulation and low intra- and inter-plate variability, making this method ideal to screen arrayed compound libraries in a 96-well format. As such, our experimental pipeline provides the proof of concept to design small screenings of PTM regulators by improving the quantitative accuracy and throughput capacity of classical western blots.Fil: Villafañez, Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina. Universidad Nacional de Córdoba; ArgentinaFil: Gottifredi, Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Soria, Ramiro Gaston. Universidad Nacional de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentin

Proposal for SPS beam time for the baby MIND and TASD neutrino detector prototypes

The design, construction and testing of neutrino detector prototypes at CERN are ongoing activities. This document reports on the design of solid state baby MIND and TASD detector prototypes and outlines requirements for a test beam at CERN to test these, tentatively planned on the H8 beamline in the North Area, which is equipped with a large aperture magnet. The current proposal is submitted to be considered in light of the recently approved projects related to neutrino activities with the SPS in the North Area in the medium term 2015-2020

Cell Production System Design: A Literature Review

Purpose In a cell production system, a number of machines that differ in function are housed in the same cell. The task of these cells is to complete operations on similar parts that are in the same group. Determining the family of machine parts and cells is one of the major design problems of production cells. Cell production system design methods include clustering, graph theory, artificial intelligence, meta-heuristic, simulation, mathematical programming. This article discusses the operation of methods and research in the field of cell production system design. Methodology: To examine these methods, from 187 articles published in this field by authoritative scientific sources, based on the year of publication and the number of restrictions considered and close to reality, which are searched using the keywords of these restrictions and among them articles Various aspects of production and design problems, such as considering machine costs and cell size and process routing, have been selected simultaneously. Findings: Finally, the distribution diagram of the use of these methods and the limitations considered by their researchers, shows the use and efficiency of each of these methods. By examining them, more efficient and efficient design fields of this type of production system can be identified. Originality/Value: In this article, the literature on cell production system from 1972 to 2021 has been reviewed

A sustainable village phone model to serve the rural developing world

Wireless technologies have created an unprecedented opportunity for rural customers in the developing world to solve their communication and information problems in an instantaneous, interactive and customized way. The framework of the study focuses on existing mobile village phone model in Bangladesh and suggests ways to make it sustainable through mobile information services marketing. The study has treated ‘village phone’ as a cost effective and interactive channel through which various time befitting information can be marketed to serve customers in the rural settings

Microfluidic Technologies for Synthetic Biology

Microfluidic technologies have shown powerful abilities for reducing cost, time, and labor, and at the same time, for increasing accuracy, throughput, and performance in the analysis of biological and biochemical samples compared with the conventional, macroscale instruments. Synthetic biology is an emerging field of biology and has drawn much attraction due to its potential to create novel, functional biological parts and systems for special purposes. Since it is believed that the development of synthetic biology can be accelerated through the use of microfluidic technology, in this review work we focus our discussion on the latest microfluidic technologies that can provide unprecedented means in synthetic biology for dynamic profiling of gene expression/regulation with high resolution, highly sensitive on-chip and off-chip detection of metabolites, and whole-cell analysis

Avian Resistance to Campylobacter jejuni Colonization Is Associated with an Intestinal Immunogene Expression Signature Identified by mRNA Sequencing

peer-reviewedThis research was funded by the The Irish Department of Agriculture and Food’s Food Institutional Research Measure (http://www.agriculture.gov.ie/ research/foodinstitutionalresearchmeasurefirm) – Grant No: 06_RDD_486.Campylobacter jejuni is the most common cause of human bacterial gastroenteritis and is associated with several post-infectious manifestations, including onset of the autoimmune neuropathy Guillain-Barré syndrome, causing significant morbidity and mortality. Poorly-cooked chicken meat is the most frequent source of infection as C. jejuni colonizes the avian intestine in a commensal relationship. However, not all chickens are equally colonized and resistance seems to be genetically determined. We hypothesize that differences in immune response may contribute to variation in colonization levels between susceptible and resistant birds. Using high-throughput sequencing in an avian infection model, we investigate gene expression associated with resistance or susceptibility to colonization of the gastrointestinal tract with C. jejuni and find that gut related immune mechanisms are critical for regulating colonization. Amongst a single population of 300 4-week old chickens, there was clear segregation in levels of C. jejuni colonization 48 hours post-exposure. RNAseq analysis of caecal tissue from 14 C. jejuni-susceptible and 14 C. jejuni-resistant birds generated over 363 million short mRNA sequences which were investigated to identify 219 differentially expressed genes. Significantly higher expression of genes involved in the innate immune response, cytokine signaling, B cell and T cell activation and immunoglobulin production, as well as the renin-angiotensin system was observed in resistant birds, suggesting an early active immune response to C. jejuni. Lower expression of these genes in colonized birds suggests suppression or inhibition of a clearing immune response thus facilitating commensal colonization and generating vectors for zoonotic transmission. This study describes biological processes regulating C. jejuni colonization of the avian intestine and gives insight into the differential immune mechanisms incited in response to commensal bacteria in general within vertebrate populations. The results reported here illustrate how an exaggerated immune response may be elicited in a subset of the population, which alters host-microbe interactions and inhibits the commensal state, therefore having wider relevance with regard to inflammatory and autoimmune disease