GSM9036721: 700 treadwear grades tire wear particles leachate treated group 2; Danio rerio; RNA-Seq
For each transcriptomic analysis, 80 zebrafish larvae were pooled per sample, with three biological replicates prepared per experimental group. Larvae were rinsed three times with PBS and preserved in RNAlater™ Stabilization Solution (Thermo Fisher) at -20°C until RNA extraction. Total RNA was isolated using a RNeasy Mini Kit (Qiagen, Hilden, Germany), and its concentration was determined with the Quant-iT™ RiboGreen® RNA Assay Kit (Invitrogen, Carlsbad, CA, USA). RNA integrity was evaluated by the Agilent TapeStation system with RNA ScreenTape (#5067-5576, Agilent Technologies). RNA samples possessing an RNA integrity number (RIN) exceeding 7.0 were utilized for library construction. For each sample, 0.5 μg of total RNA was used to create libraries using a commercial kit (TruSeq Stranded Total RNA Library Prep Gold Kit; Illumina, San Diego, CA, USA). Ribosomal RNA (rRNA) was initially depleted from the total RNA. Isolated mRNAs were fragmented into short pieces, and double-stranded cDNAs were synthesized. The cDNAs were modified through end-repair, poly A addition, and connected with sequencing adapters. Library concentration was measured using the KAPA Library Quantification Kit for Illumina platforms (KAPA Biosystems, Wilmington, MA, USA), and the library size distribution was evaluated with the Agilent D1000 ScreenTape (#5067-5582, Agilent Technologies).