1. Genome-scale RNAi screens for high-throughput phenotyping in bloodstream-form African trypanosomes.
- Author
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Glover L, Alsford S, Baker N, Turner DJ, Sanchez-Flores A, Hutchinson S, Hertz-Fowler C, Berriman M, and Horn D
- Subjects
- Africa, Gene Library, Genetic Testing methods, Genome, Protozoan genetics, Phenotype, RNA Interference, Trypanosoma genetics
- Abstract
The ability to simultaneously assess every gene in a genome for a role in a particular process has obvious appeal. This protocol describes how to perform genome-scale RNAi library screens in bloodstream-form African trypanosomes, a family of parasites that causes lethal human and animal diseases and also serves as a model for studies on basic aspects of eukaryotic biology and evolution. We discuss strain assembly, screen design and implementation, the RNAi target sequencing approach and hit validation, and we provide a step-by-step protocol. A screen can yield from one to thousands of 'hits' associated with the phenotype of interest. The screening protocol itself takes 2 weeks or less to be completed, and high-throughput sequencing may also be completed within weeks. Pre- and post-screen strain assembly, validation and follow-up can take several months, depending on the type of screen and the number of hits analyzed.
- Published
- 2015
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