title: Transcriptome analysis of amphiprion sebae brain2 sample
design: Library preparation was performed following Illumina TruSeq RNA library protocol outlined in “TruSeq RNA Sample Preparation Guide” . 1ug of Total RNA was subjected to Poly A purification of mRNA. Purified mRNA was fragmented for 2 minutes at elevated temperature (94oC) in the presence of divalent cations and reverse transcribed with Superscript III Reverse transcriptase by priming with Random Hexamers. Second strand cDNA was synthesized in the presence of DNA Polymerase I and RnaseH. The cDNA was cleaned up using HighPrep PCR (MAGBIO, Cat# AC-60050). Illumina Adapters were ligated to the cDNA molecules after end repair and addition of A base. SPRI cleanup was performed after ligation. The library was amplified using 8 cycles of PCR for enrichment of adapter ligated fragments. The prepared library was quantified using Nanodrop and validated for quality by running an aliquot on High Sensitivity Bioanalyzer Chip (Agilent).