For calculating expression levels from RNA-Seq coverage data, the uniquely transcribed region(s) for each localized gene was determined by taking regions covered by exons of the gene and excluding transcribed regions from any overlapping genes, both with respect to genes lying on same strand (for calculation using strand-specific RNA-Seq coverage data), and for genes on either strand (for calculation using unstranded RNA-Seq coverage data). RNA-Seq coverage read-count data was then correlated by location with the uniquely transcribed region(s) of each gene to produce the sum of reads over the entire uniquely transcribed region for the gene. Reads per kilobase of exon model per million mapped reads (RPKM) was then calculated using the method from Motazavi et al, Nat. Methods 6, 621-628 (2008). (RPKM = 10^9 * C / N * L * R, where C = number of reads in gene, N = number of uniquely mappable reads in the experiment, L = sum of uniquely transcribed bases in bp, and R = read length in bp). RPKM levels and expression level bins are recalculated for a gene whenever modifications are introducted to one or more transcripts in its genomic region.
The RPKM values were further binned into eight expression level bins (by RPKM value) to improve querying capabilities and displays.
Bin 1: No/Extremely low expression (0)
Bin 2: Very low expression (1-9), percentiles 1-25, approximately.
Bin 3: Low expression (10-150), percentiles 26-50, approximately.
Bin 4: Moderate expression (151-2500), percentiles 51-75, approximately.
Bin 5: Moderately high expression (2501-7500), percentiles 76-85, approximately.
Bin 6: High expression (7501-25000), percentiles 86-95, approximately.
Bin 7: Very high expression (25001-100000), percentiles 96-99, approximately.
Bin 8: Extremely high (>100000), the 100th percentile, approximately.
For calculating expression levels from the FlyAtlas larval and adult tissue data, FlyAtlas provided mean coverage levels for Affy2 probe sets, and FlyBase intersected these probe sets with the exons of all FlyBase genes (using D. melanogaster Annotation Release 5.26) to associate each probe set with one or more FlyBase genes. These values were distributed into five expression level bins for further analysis.
TO DOWNLOAD THESE DATA: Bulk data files can be accessed from the Precomputed Data Files page: from the top menu button 'Files', select 'Current Release'; open the 'Genes' section; the item line is 'RNA-Seq RPKM values'.