Transcriptomic analysis of the liver from control sibling and mutant uhrf1-hi272 zebrafish larvae at 120 hpf treated with DMSO or Palbociclib (PD)

This study was carried out to compare the changes in gene expression in livers of 120 hpf control sibling and mutant uhrf1-hi272 zebrafish larvae in response to treatment with DMSO or Palbociclib (PD) 48 hpf embryos from an incross of uhrf1-hi272+/- zebrafish adults with Tg(fabp10a:CAAX-EGFP) to mark the livers were treated with either 0.5% DMSO or 20 microM PD in 0.5% DMSO in 6 well plates. At 120 hpf, 15- 20 livers from were dissected from larvae treated with PD or DMSO treated and individual livers were immediately placed in TRIzol and stored at -80 while the carcass of each larvae was genotyped. The individual livers from uhrf1hi272-/- mutants and from WT in each treatment batch were pooled RNA was extracted per standard TRIzol/Chloroform method and concentrated, DNAse 1 treated and resuspended in DNase/RNase free water. For library preparation, high quality RNA was identified based on bioanalyzer analysis and quantified so that 100 ng of total RNA was used to initiate the library preparation using Ribo-Zero rRNA Removal Kit (Human/Mouse/Rat). The RNA sample was chemically fragmented and random primed for reverse transcription using Illumina TruSeq V2 RNA sample Prep Kit and first-strand cDNA reverse transcribed using random primers. Following second-strand cDNA synthesis, and end repair. TrueSeq adapter-index was ligated to cDNA libraries, and PCR amplification of 12 cycles was done for enrichment, producing a 350-400 bp fragment including adapters. The fragment sizes and purity of the libraries were confirmed by analyzing on a Bioanalyzer 2100 and were sequenced using the Illumina NextSeq 550 system with a Paired-end HO V2.5 kit, 150 cycles.