Open Access Research article

Advances in genome-wide RNAi cellular screens: a case study using the Drosophila JAK/STAT pathway

Katherine H Fisher2, Victoria M Wright2, Amy Taylor1, Martin P Zeidler2* and Stephen Brown1*

Author Affiliations

1 The Sheffield RNAi Screening Facility, Department of Biomedical Science, University of Sheffield, Alfred Denny Building, Western Bank, Sheffield S10 2TN, UK

2 The MRC Centre for Developmental and Biomedical Genetics and The Department of Biomedical Science, University of Sheffield, Firth Court, Western Bank, Sheffield S10 2TN, UK

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BMC Genomics 2012, 13:506  doi:10.1186/1471-2164-13-506

Published: 24 September 2012

Additional files

Additional file 1:

Plate layouts of HFA and SRSFv1 libraries. (A) Layout of HFA library plates including the position of positive (blue) and negative pathway regulators, used as controls in the Müller et al. screen. (B) Layout of SRSFv1 library including the DIAP1 barcode (black), technical controls (purple), non-interacting controls (yellow) and positive (blue) and negative (red) pathway regulators used as controls.

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Additional file 2:

Quality control of HFA and SRSF JAK/STAT screens. Box and whisker plots representing each plate from separate replicates in HFA and SRSF screens. Asterisks denote plates where variance can be observed by eye.

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Additional file 3:

dsRNAs resulting in high RL values can give skewed FL/RL ratios. List of genes highlighted in red circle in Figure 3D have high RL values, many of which have unaffected FL values. All values are robust Z-scores averaged over three replicates.

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Additional file 4:

Hits identified in the SRSF screen are also present in the HFA collection. List of hits identified in SRSF screen, as shown in Table 2, but including dsRNA amplicon names for SRSF and HFA libraries, as well as Flybase IDs.

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Additional file 5:

Tertiary screening results and primer design. List of the hits identified in both SRSF and HFA screens. Included are Z-scores calculated from FL/RL ratios and averaged across triplicates, for genome and tertiary screens. Significance >2 or <−2 highlighted in grey and bold, significance >1.7 or <-1.7 highlighted in blue and mark #. All sequence information is included where novel designs were made. Some genes were not screened (NS) in the tertiary screen either due to inability to target independent regions, independent regions had already been screened, or the gene is now withdrawn from Flybase.

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Additional file 6:

The 6 genes found to be significant in both SRSF and Baeg screens. Fold change values are shown as originally presented in ref [15], and +/- indicates an increase or decrease in reporter activity, respectively. Grey/blue boxes highlight significance levels as indicated in key.

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