Poly a selection kit

RNA from embryos was extracted using TriZol (Thermo Fisher Scientific Cat No 15596026) according to manufacturer protocol. 1.25ug of Total RNA was submitted for RNA-Sequencing to the NCI CCR Genomics Core. The libraries were made using Poly(A) Selection kit (illumina). 150bp fragments were sequenced by paired-end 75bp utilizing Illumina NextSeq. Reads were aligned to Ensembl Zebrafish Genome Build 11, GRCz11, using Histat21. SAMtools was used for sorting and indexing2. PartekFlow was used for ANOVA and pathway analysis.

Total RNA was prepared using the Sigma GenElute total mammalian RNA miniprep kit with optional DNase step, according to the protocol of the manufacturer. RNA quality was assessed by Nanodrop and an Agilent RNA Screen Tape System, and 1ug was used for library preparation using RNA with Poly A selection kit (Illumina), as per the manufacturer’s instructions. Sequencing was performed on an Illumina HiSeq. RNA-sea FASTQ data were processed and mapped to the mouse reference genome (GRCm38) using CLC Genomics Workbench 20 (Qiagen). Differential gene expression was performed using the DESeq2 package in R [43 (link)]. Heatmaps were made in R using the pheatmap: pretty heatmaps package shown as the log₂RPKM. Raw sequencing files have been deposited in Sequence Read Archives (SUB8204864, PRJNA665496).

Total RNA was prepared using the Sigma GenElute total mammalian RNA miniprep kit with an optional DNase step, according to the protocol of the manufacturer. RNA quality was assessed by a Nanodrop instrument and an Agilent RNA Screen Tape system, and 1 μg was used for library preparation using RNA with a poly(A) selection kit (Illumina), according to the manufacturer’s instructions. Sequencing was performed on an Illumina HiSeq platform. RNASeq FASTQ data were processed and mapped to the mouse reference genome (GRCm38) using CLC Genomics Workbench 20 (Qiagen). Differential gene expression was determined using the DESeq2 package in R (40 (link)). Heatmaps and volcano plots were made in GraphPad Prism 9. Raw sequencing data were deposited under BioProject PRJNA838604.