Exclusion of non-expressed genes from the Chikungunya RNAi screen hit identification process

To allow exclusion of non-expressed genes from the hit identification process, microarray analyses of (i) Allstars transfected and Chikungunya (CHIKV) infected, (ii) Allstars transfected and non-infected, (iii) non-transfected and CHIKV infected and (iv) non-transfected and non-infected 293 cells were performed. Transfection was done by adding 4x10e5 293 cells (in complete DMEM medium) to a mixture of serum-free RPMI medium, HiperFect (Qiagen) and Allstars siRNAs (Qiagen, final siRNA concentration: 20 nM). Infection with CHIKV-GFP was performed 72 h p.t. at an MOI of ~3. Cells were lysed 18 h p.i. using Trizol (Invitrogen), followed by RNA isolation, that was subsequently subjected to microarray analysis. Microarray experiments were performed as dual-color hybridizations on Agilent 4x44K human whole genome catalog arrays (Agilent-014850). To compensate for dye-specific effects, a dye-reversal color-swap was applied.