title: RNA-Seq of C. elegans: eat-2(-)_control RNAi_Day 1_01
design: L1 synchronized worms {WT or eat-2(ad1116)} were grown on RNAi-seeded plates till L4/YA, after which FUDR was added to prevent progeny growth. After about 24 hours, worms were collected in M9 buffer and washed thrice and then frozen in Trizol (Invitrogen, USA). The worms were collected, passed through a freeze-thaw cycle and lysed by vigorous vortexing. RNA was purified by phenol:chloroform:isoamylalcohol extraction followed by ethanol precipitation.  RNA were isolated as mentioned above. Quality Control of RNA was done using Bioanalyzer 2100 RNA 6000 NanoAssay chip and RNA above RIN = 8 was included to the study (Agilent Technologies, Santa Clara, USA).  cDNA library was subsequently constructed by TruSeq RNA Library Prep Kit v2 (Illumina). Sequencing (34bp single end) was performed with two biological replicates for different treatments using Illumina Genome Analyzer IIx (GA IIx). Imaging, base calling and quality scoring were performed as per standard manufacturers guidelines (Illumina, USA).The de-multiplexing and conversion of BCL file format reads to FASTQ file format was done with Illumina-supported CASAVA v1.8.2 software package.