Identifying suitable experimental designs to determining differentially expressed genes and benchmark doses from transcriptomic dose-response studies in zebrafish embryos
To identify suitable experimental designsfor toxicology experiments (i.e., sample size, pool size, number of biological replicates, and number of doses), we exposed zebrafish embryos to 0.1% dimethyl sulfoxide for 4-120 hours post fertilization. We then created 96 samples to examine the impact of pool size comprised of 1, 5, 10, and 20 zebrafish embryos each (n = 24). From these, total RNA extraction was extracted and transcriptomic analyses were done using the TempO-Seq S1500+ Surrogate Assay platform. Samples were sequencing on an Illumina NextSeq 2000. The data were simulated to conduct power analyses exploring suitable study designs for identification of differentially expressed genes and for benchmark dose analysis. The aim was to identify a feasible pool size and sample size that would yield an acceptable false discovery rate for these analyses. Overall design: Samples were comprised of different pool sizes of embryos (i.e., 1, 5, 10, and 20 embryos per sample, indicated by Z1, Z5, Z10, and Z20) to identify optimal experimental designs for the dose-response study. Each pool size included 24 samples and a total of 96 samples were sequenced on the TempO-Seq S1500+ Surrogate assay platform. This study consisted of 12 extraction batches with 8 samples per batch and 3 batches per pool size.