Project description:Purpose: The goal of this study was to compare 3’ RNA-seq data to standard sequencing in samples collected from zebrafish exposed to control conditions or an elevated level of perfluorobutane sulfonamide (FBSA). Methods: Dechorionated embryonic zebrafish were statically exposed at 8 hours post fertilization (hpf) to either vehicle control (0.5% methanol) or to 47 µM perfluorobutane sulfonamide (FBSA) in 96-well plate format. Zebrafish embryos were sampled at 48 hpf, before morphological effects were observable but with samples phenotypically anchored to abnormal morphology at 120 hpf. At 48 hpf, 10 embryos were pooled per replicate (n = 3), euthanized via ice bath, and total RNA extracted using a Zymo Research Direct-zol RNA MiniPrep kit. For the standard RNA-seq, library preparation and sequencing was performed by Beijing Genomics Institute (BGI) on their BGISEQ-500 platform (raw data files previously submitted to GEO; GSE186576). For the 3' RNA-seq, library preparation and sequencing was performed by the Center for Quantitative Life Sciences (CQLS) at Oregon State University using the Lexogen QuantSeq 3' FWD library preparation kit and a HiSeq 3000. Both datasets were analyzed using the same RNA-seq analysis pipeline. Results: We found that 3’ RNA-seq and standard showed specific alignment advantages when focusing on annotated or unannotated regions of the genome, with 3’ RNA-seq showing better alignment to annotated regions. We also found that 3’ RNA-seq was far better at detecting DEGs under conditions of sparse data (fewer reads) but that DEGs identified by standard RNA-seq led to a greater number of statistically enriched biological functions.
