sterechinus neumayeri transcriptome
Total RNA was extracted from 13 pooled larval samples detailed in Table 1. Each sample included 5000 larvae, except sample 9 (Day 30, μatm 1071) which included 3500 due to sample limitations. Further RNA extraction details are found in the Appendix S1 (Supporting information). A yield of 13 μg of total RNA equally representing each sample was combined into a single pool for the construction of a cDNA library. The cDNA library was built at the Center for Genomics and Bioinformatics (CGB) at Indiana University, using a SMART cDNA synthesis kit (Clontech, Mountain View, CA) via manufacturer instructions and previously published protocols (Meyer et al. 2009; Schwartz et al. 2010; Der et al. 2011). The cDNA was normalized using a customized protocol developed and published by the CGB specifically designed for 454 sequencing (Lai et al. 2012). These methods ensure polyadenylated (polyA) transcript selection and gene copy normalization to identify and sequence low-copy transcripts, maximizing transcript diversity