RNA-Seq of danio rerio: whole organism embryo PFOA (CAS 45285-51-6)
At the end of each PFAS exposure period, zebrafih embryos were homogenized in Qiagen buffer RLT and betamercaptoethanol, then frozen. RNA was isolated with Ambion MagMAX-96 Total RNA Isolation kit. Samples were transferred to 0.5 mL Eppendorf tubes in prepration for RNA-Seq library prepation. RNA was normalized to 6 ng/uL using epMotion 5075. Illumina-compatible Lexogen CORALL RNA-Seq libraries were then prepared for RNA sequencing. Library pools were quality-checked and quantified using a combination of Qubit dsDNA HS, Agilent 4200 TapeStation HS DNA1000 and Invitrogen Collibri Library Quantification qPCR assays by the Michigan State University RTSF Genomics Core. Each pool was loaded onto 1 lane of a NovaSeq S4 flow cell. Sequencing was performed by Michigan State University in a 2x150bp paired end format using a NovaSeq 6000 v1.5 300 cycle reagent kit. Base calling was done by Illumina Real Time Analysis (RTA) v3.4.4 and output of RTA was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v2.20.0.