Installation instructions

Download the extension to your home directory/GENE-E/extensions

RIGER

RIGER-E may be downloaded and used free of charge by academic and other non-profit researchers. Commercial users please contact us for licensing terms. Download

RIGER ranks shRNAs according to their differential effects between two classes of samples, then identifies the genes targeted by the shRNAs at the top of the list. In this way, RIGER identifies genes essential to the difference between the classes. For details, see Luo, Cheung, Subramanian, et al. (2008).

1. The Class Comparisons parameter defines one or more class comparisons on which to run RIGER. Click Edit to open the Edit Comparison window. You can select columns by categories or by name. For each class comparison, you select columns for the two classes to be compared: Highlight the desired categories (or columns) in the middle panel. Click an arrow icon to move them to class A or B. To remove categories (or columns) from a class, highlight them and click the delete icon. GENE-E moves them back to the middle panel.
2. Optionally add another two class comparison by clicking the add button.
3. Select a method to analyze your RNAi screening data.

Second Best Rank
A method based on ranking genes by the rank of the second best scoring hairpin for that gene. This is currently the preferred method for RNAi screening analysis in the RNAi Platform.
Kolmogorov-Smirnov
A statistical method and the basis for the original RIGER as described in Luo et al, PNAS, 2008
Weighted Sum
This method is a modification of the Second Best Rank in that it takes the combined sum of the first and second best ranks for hairpins for a given gene. The best ranking hairpin is given a weight of 0.25 and the second best ranking hairpin is given a weight of 0.75. The sum of these weighted ranks is used to compute a new score, and genes are ranked by this new score.

Regardless of the method you choose to use, you may use either Signal to Noise or Log Fold Change to generate a single hairpin level score from the set of replicates for each hairpin in your screen. All the methods, including Kolmogorov-Smirnov, can use either Signal to Noise or Log Fold Change. You may also use the T-Test option, although this is still experimental and has not been fully tested.

Please note that we now compute the FDR as a p-value, and therefore additional permutations are required for the null distribution.

When it is finished, RIGER results are listed in the Navigator window as shown below. Double-click the 'NES' or 'Scored shRNAs' to view the results.

Scored shRNAs
Double-click 'Scored shRNAs' to display a heat map showing the ranked list of shRNAs and their scores. The heat map is sorted by the scores of the first comparison. Select Tools>Sort Rows to sort a different comparison by score.

NES: Double-click 'NES' to display a heat map showing the normalized enrichment scores (NES) for the genes in your dataset. The heat map is sorted by the NES of the first comparison. Select Tools>Sort Rows to sort a different comparison by the NES.

Analysis Report
To view a report of RIGER results, right-click either RIGER result node and select View Gene Report or View Hairpin Report. To save the report, right-click either RIGER result node and select Save Gene Report or Save Hairpin Report. The report lists: Gene, Hairpins, # Hairpins, # Hairpins in the top 500, 1000, 5000, and 10000, NES, Gene rank, p-value, and p-value rank.