mRNA sequencing in cde-1 mutants versus WT C. elegans infected by the Orsay virus

Aim: We question relative mRNA levels in cde-1 mutants versus WT C. elegans (N2 strain) infected by the Orsay virus Methods: Approximately 200 cde-1 (tm1021) mutants or N2 animals were infected for 48 hours from the L2 stage to the adult stage with 20 µl filtrate of the Orsay virus on 50mm plate seeded with HB101 bacteria, in biological triplicates. Animals were then washed in M9 and resuspended in 1 ml TRIsure (Bioline). Samples were freeze-thaw three times using liquid nitrogen and RNA purification was performed according to Bioline's guidelines. mRNA libraries were prepared using the NEBNext Ultra directional RNA library kit (NEB) with poly(A) selection and deep sequencing was produced with an Illumina HiSeq machine (single read 50). For differential expression analysis, genes were only included in the analysis if they had at least 1 count per million in all of the control samples or all of the experiment samples. After applying this filter, differentially expressed genes were then called using EdgeR (Robinson, et al. 2010). For the correlation matrix, Pairwise Pearson correlation values were calculated between all samples. Overall design: Examination of mRNA profiles in Orsay virus infected animals from two different genetic backgrounds