GSM8640474: Spc cells, WT, rep1; Mus musculus; RNA-Seq
Sorted germ cells of littermates were collected and frozen in Trizol until triplicates of each genotype were collected. RNA extraction was performed using the DirectZol RNA MicroPrep kit (Zymo, R2061). NGS library prep was performed with Illumina's Stranded mRNA Prep Ligation Kit following Stranded mRNA Prep Ligation ReferenceGuide (April 2021) (Document 1000000124518 v02). Libraries were prepared with a starting amount of 10 ng and amplified in 13 PCR cycles. Two post PCR purification steps were performed to exclude residual primer and adapter dimers. Libraries were profiled in a DNA High Sensitivity chip on a 2100 Bioanalyzer (Agilent technologies) and quantified using the Qubit 1x dsDNA HS Assay Kit, in a Qubit 4.0 Fluorometer (Invitrogen by Thermo Fisher Scientific).  All samples were pooled together in equimolar ratio aiming for at least 50M reads per sample and sequenced on a NextSeq 2000 P3 (200 cycles) FC, PE for 2x108 cycles plus 2x10 cycles for the index reads and 1 dark cycle upfront R1 and R2.