High-Throughput Expression Screening in Mammalian Suspension Cells

2020 ◽  
pp. 117-125
Author(s):  
Susan D. Chapple ◽  
Michael R. Dyson
Keyword(s):  
High Throughput ◽  
Suspension Cells ◽  
Expression Screening

scholarly journals Construction of a series of vectors for high throughput cloning and expression screening of membrane proteins from Mycobacterium tuberculosis

2008 ◽  
Vol 8 (1) ◽  
pp. 51 ◽  
Author(s):  
Huajun Qin ◽  
Jian Hu ◽  
Yuanzhi Hua ◽  
Shridhar V Challa ◽  
Timothy A Cross ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Membrane Proteins ◽  
High Throughput ◽  
Cloning And Expression ◽  
Expression Screening

High-throughput protein expression screening and purification in Escherichia coli

Methods ◽  
2011 ◽  
Vol 55 (1) ◽  
pp. 65-72 ◽  
Author(s):  
Renaud Vincentelli ◽  
Agnès Cimino ◽  
Arie Geerlof ◽  
Atsushi Kubo ◽  
Yutaka Satou ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Protein Expression ◽  
High Throughput ◽  
Expression Screening

scholarly journals A high-throughput expression screening platform to optimize the production of antimicrobial peptides

2017 ◽  
Vol 16 (1) ◽  
Author(s):  
Christine Schreiber ◽  
Hagen Müller ◽  
Oliver Birrenbach ◽  
Moritz Klein ◽  
Doreen Heerd ◽  
...  
Keyword(s):  
Antimicrobial Peptides ◽  
High Throughput ◽  
Expression Screening ◽  
Screening Platform

scholarly journals A Cell-Based High-Throughput Screening Assay for Posttranscriptional Utrophin Upregulation

2012 ◽  
Vol 18 (4) ◽  
pp. 400-406 ◽  
Author(s):  
Catherine Moorwood ◽  
Neha Soni ◽  
Gopal Patel ◽  
Steve D. Wilton ◽  
Tejvir S. Khurana
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Posttranscriptional Regulation ◽  
Therapeutic Potential ◽  
Dystrophin Gene ◽  
Screening Assay ◽  
Expression Screening ◽  
High Throughput Screening Assay ◽  
Promising Therapeutic Approach ◽  
Muscle Wasting Disease

Duchenne muscular dystrophy (DMD) is a devastating muscle-wasting disease caused by mutations in the dystrophin gene. Utrophin is a homologue of dystrophin that can compensate for its absence when overexpressed in DMD animal models. Utrophin upregulation is therefore a promising therapeutic approach for DMD. Utrophin is regulated at both transcriptional and posttranscriptional levels. Transcriptional regulation has been studied extensively, and assays have been described for the identification of utrophin promoter-targeting molecules. However, despite the profound impact that posttranscriptional regulation has on utrophin expression, screening assays have not yet been described that could be used to discover pharmaceuticals targeting this key phase of regulation. We describe the development and validation of a muscle cell line–based assay in which a stably expressed luciferase coding sequence is flanked by the utrophin 5′- and 3′-untranslated regions (UTRs). The assay was validated using the posttranscriptional regulation of utrophin by miR-206. The assay has a Z′ of 0.7, indicating robust performance in high-throughput format. This assay can be used to study utrophin regulatory mechanisms or to screen chemical libraries for compounds that upregulate utrophin posttranscriptionally via its UTRs. Compounds identified via this assay, used alone or in a synergistic combination with utrophin promoter-targeting molecules, would be predicted to have therapeutic potential for DMD.

scholarly journals A high-throughput RNA-seq approach to profile transcriptional responses

2015 ◽  
Author(s):  
Gregory A Moyerbrailean ◽  
Gordon O Davis ◽  
Chris T Harvey ◽  
Donovan Watza ◽  
Xiaoquan Wen ◽  
...  
Keyword(s):  
Gene Expression ◽  
High Throughput ◽  
Deep Sequencing ◽  
Transcriptome Profiling ◽  
Iterative Refinement ◽  
Specific Gene ◽  
Rna Seq ◽  
Transcriptional Responses ◽  
Expression Screening

In recent years, different technologies have been used to measure genome-wide gene expression levels and to study the transcriptome across many types of tissues and in response to in vitro treatments. However, a full understanding of gene regulation in any given cellular and environmental context combination is still missing. This is partly because analyzing tissue/environment-specific gene expression generally implies screening a large number of cellular conditions and samples, without prior knowledge of which conditions are most informative (e.g. some cell types may not respond to certain treatments). To circumvent these challenges, we have established a new two-step high-throughput and cost-effective RNA-seq approach: the first step consists of gene expression screening of a large number of conditions, while the second step focuses on deep sequencing of the most relevant conditions (e.g. largest number of differentially expressed genes). This study design allows for a fast and economical screen in step one, with a more profitable allocation of resources for the deep sequencing of re-pooled libraries in step two. We have applied this approach to study the response to 26 treatments in three lymphoblastoid cell line samples and we show that it is applicable for other high-throughput transcriptome profiling requiring iterative refinement or screening.

scholarly journals A sample preparation protocol for high throughput immunofluorescence of suspension cells

2020 ◽  
Author(s):  
Anna Bäckström ◽  
Laura Kugel ◽  
Christian Gnann ◽  
Hao Xu ◽  
Joseph E. Aslan ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Sample Preparation ◽  
High Throughput ◽  
Spatial Information ◽  
Protein Localization ◽  
Cell Types ◽  
Human Platelets ◽  
Adherent Cell ◽  
Suspension Cells ◽  
High Throughput Imaging

AbstractImaging is a powerful approach for studying protein expression and has the advantage over other methodologies in providing spatial information in situ at single cell level. Using immunofluorescence and confocal microscopy, detailed information of subcellular distribution of proteins can be obtained. While adherent cells of different tissue origin are relatively easy to prepare for imaging applications, nonadherent cells from hematopoietic origin, present a challenge due to their poor attachment to surfaces and subsequent loss of a substantial fraction of the cells. Still, these cell types represent an important part of the human proteome and express genes that are not expressed in adherent cell types. In the era of cell mapping efforts, overcoming the challenge with suspension cells for imaging applications would enable systematic profiling of hematopoietic cells. In this work, we successfully established an immunofluorescence protocol for preparation of suspension cell lines and peripheral blood mononucleated cells (PBMC) and human platelets. The protocol is based on a multi-well plate format with automated sample preparation, allowing for robust high throughput imaging applications. In combination with confocal microscopy, the protocol enables systematic exploration of protein localization to all major subcellular structures.

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