Next Article in Journal / Special Issue
RNAi in Arthropods: Insight into the Machinery and Applications for Understanding the Pathogen-Vector Interface
Previous Article in Journal / Special Issue
The Influence of Competition Among C. elegans Small RNA Pathways on Development
Genes 2012, 3(4), 686-701; doi:10.3390/genes3040686
Review

Using Multiple Phenotype Assays and Epistasis Testing to Enhance the Reliability of RNAi Screening and Identify Regulators of Muscle Protein Degradation

1,* , 2
, 3
 and 1,*
1 Medical Resarch Council/Arthritis Research UK Centre for Musculoskeletal Ageing Research, University of Nottingham, Royal Derby Hospital, Derby, DE22 3DT, UK 2 Medical Resarch Council/Arthritis Research UK Centre for Musculoskeletal Ageing Research, University of Nottingham, School of Veterinary Medicine & Science, Sutton Bonington, LE12 5RD, UK 3 Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA
* Authors to whom correspondence should be addressed.
Received: 13 August 2012 / Revised: 27 September 2012 / Accepted: 30 October 2012 / Published: 2 November 2012
(This article belongs to the Special Issue RNA Interference)
View Full-Text   |   Download PDF [416 KB, 5 November 2012; original version 2 November 2012]   |   Browse Figures

Abstract

RNAi is a convenient, widely used tool for screening for genes of interest. We have recently used this technology to screen roughly 750 candidate genes, in C. elegans, for potential roles in regulating muscle protein degradation in vivo. To maximize confidence and assess reproducibility, we have only used previously validated RNAi constructs and have included time courses and replicates. To maximize mechanistic understanding, we have examined multiple sub-cellular phenotypes in multiple compartments in muscle. We have also tested knockdowns of putative regulators of degradation in the context of mutations or drugs that were previously shown to inhibit protein degradation by diverse mechanisms. Here we discuss how assaying multiple phenotypes, multiplexing RNAi screens with use of mutations and drugs, and use of bioinformatics can provide more data on rates of potential false positives and negatives as well as more mechanistic insight than simple RNAi screening.
Keywords: RNAi; systems biology; network biology; functional genomics; muscle; proteolysis; C. elegans RNAi; systems biology; network biology; functional genomics; muscle; proteolysis; C. elegans
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Share & Cite This Article

Export to BibTeX |
EndNote


MDPI and ACS Style

Lehmann, S.; Shephard, F.; Jacobson, L.A.; Szewczyk, N.J. Using Multiple Phenotype Assays and Epistasis Testing to Enhance the Reliability of RNAi Screening and Identify Regulators of Muscle Protein Degradation. Genes 2012, 3, 686-701.

View more citation formats

Related Articles

Article Metrics

Comments

Citing Articles

[Return to top]
Genes EISSN 2073-4425 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert