Since many reports have also used a window that flanks the TSS of known genes [ 16, 17], we have also defined a different window of same size (5 KB) now encompassing 2,5 KB in each side of a given TSS.
A gene is termed as uni-directional if no oppositely oriented TSS is found within 10 kb upstream of the given TSS, or if a similarly oriented TSS is found at least 1 kb upstream.
First and foremost, Cuffdiff2 is designed specifically to test differential isoform usage for a given transcription start site (TSS).
Since the TSSs for a given gene are not fixed but vary on the chromosome, a representative TSS was defined for each gene, as described in Yamashita et al. [ 27].
Due to the widespread existence of alternative TSSs for a given gene and the expected preferred distance of LTSM relative to the TSS, we created a transcript-based sequence set from EnsEMBL55, spanning from the TSS to +200 bp downstream.
Since, for a given gene, several closely spaced TSSs can be identified (Pelechano et al., 2013), we used a peak-calling procedure to define, in three biological replicates, TSS clusters corresponding to transcript isoforms with closely spaced 5′-ends (TSSCs; see 'Materials and methods').
GAM models included a 0 1 response variable indicating whether a gene belonged to the set of psoriasis DEGs, along with two covariates (x 1 and x 2) corresponding to the total number of TSS-proximal matches for a given PWM model (x 1) and the length of non-masked sequence scanned for each gene (x 2) [25].
Transcript heterogeneity at the 5′-end is particularly relevant for NMD targeting since the use of different transcription start sites (TSSs) will generate, for a given gene, mRNAs with different 5′-UTRs that may include uORFs.
EPD considers the most frequent cDNA 5 end as the TSS and applies a specialized algorithm to discover multiple promoters for a given gene, whereas PlantProm DB contains plant promoters based on published TSS mapping data [ 27].
Kinetic constants values were finally correlated to the sludge VSS/TSS ratio (VSS = Volatile Suspended Solids; TSS = Total Suspended Solids) in order to provide a tool for a gross estimation of WO kinetic parameters, for a given sludge, relying on simple analytical measurement (VSS and TSS), commonly available at every WWTP.
