Training course at the Scottish Institute for Cell Signaling (SCILLS)
University of Dundee, Scotland
Title: Upstream Workshop on siRNA screening
Organizers: Ron Hay, Honorary Programme Leader SCILLS, r.t.hay@dundee.ac.uk
and Amit Garg, Data Manager, SCILLS, A.K.Garg@dundee.ac.uk
Dates: To be decided
Location: Sir James Black Centre, University of Dundee, Dundee, Scotland
Duration: 5 days
Topics covered: The participants will be trained in the following areas:
Fundamentals of gene silencing using siRNA (lecture).
Ablation of defined siRNA targets. Analysis of knockdown by RT.PCR and Western blotting
(practical).
Cell based assays for siRNA screening (practicals)
-Viability and synthetic lethality assays. Luciferase based ATP level determination.
Generation of home made kits for such assays.
-Transcriptional assays using cell lines containing an NF-kB dependent luciferase reporter.
Luciferase based assay. Generation of home made kits for such assays.
-High content imaging with GFP target proteins in reporter cell lines. Use of high content
microscopes and familiarization with software for data collection and analysis.
Use of liquid handling robots for siRNA screen setup (practical).
Data handling. Activity Base software for plate tracking and data analysis. Hands on computer
training.
Establishing assay robustness. Statistical analysis of screening data to determine Z’ scores (assay
robustness). Wet practical and computer based analysis.
Dealing with “off target” effects: pool deconvolution and retesting (practical); pharmacological
inhibition of identified pathways (practical); generation of siRNA resistant expression constructs
(discussion).
We will require a considerable amount of disposable material for the course, including 96 well
assay plates, tips for liquid handling robots, siRNAs, transfection reagents and tissue culture reagents..
To costs to a minimum the organizer will provide the participants with in-house made assay kits, while
the remaining reagents will be purchased. The organizers will provide all the necessary reporter cell
lines.
Estimated cost: 500€ per student. Does not include accommodations, meals (lunch at the
course provided) nor trips. Based on our commercial estimations we will need approximately
2500 EURO to cover all remaining costs. Number of participants 3-5, timing: 5 days.
Type of course: hands on workshop. Practical work 50%, data analysis 15%, seminars 15%,
demonstrations 10%, discussion panels and participant presentations 10%