Description
To obtain a global view of gene activity in prothoracic glands at single-nucleotide resolution, high-throughput RNA-Seq experiments using Illumina sequencing technology on poly (A)–enriched RNAs of 3 biological samples each from day 0 and day 6 of the 5th instar were performed.The lllumina® mRNA-Seq Sample Prep Kit was used to process the samples according to the manufacturer's instructions. mRNA was isolated from total RNA using oligo-dT magnetic beads. After fragmentation of the mRNA, cDNA synthesis was performed. This was used for ligation with the sequencing adapters and PCR amplification of the resulting products. Quality and yield after sample preparation was measured with the Advanced Analytical Fragment Analyzer. The size of the resulting products was consistent with the expected size distribution with a broad peak between 200-500 bp on a DNA 1000 chip). A concentration of 17 pM of DNA was used for clustering and DNA sequencing on lllumina cBot and HiSeq2500 (HCS v2.2.58 software).