GSM8640479: Germ cells, Dnmt3CKO/KO, Nrf1KO/KO rep1; Mus musculus; RNA-Seq
Dissected whole testis of mutants and littermate controls were collected and frozen  in Trizol until triplicates of each genotype were collected. Testes were homogenized in Trizol using an IKA T10 basic homogenizer and 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 29 ng and amplified in 12 PCR cycles. Two post PCR purification steps were performed to exclude residual primer and adapter dimers. Libraries were profiled in a High Sensitivity DNA 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 in equimolar ratio and sequenced on a NextSeq2000 P4 FC (200 cycles), PE for 2x 108 cycles plus 2x 10 cycles for the dual index read and 1 dark cycle upfront both R1 and R2.