GSM1542522: Dicer WT Pach RNA; Mus musculus; RNA-Seq
Testes from adult Dgcr8 and Dicer cKOs, together with WT control littermates (day 70-80 pp) were removed, weighed and decapsulated prior to enrichment of specific spermatogenic cell types using the STA-PUT method based on separation by cell diameter/density at unit gravity (Bellvé, 1993). Purity of resulting fractions was determined by microscopy based on cell diameter and morphology. Pachytene cells were approximately 90% pure, with residual possible contamination from spermatocytes of slightly earlier or later developmental stages. Leptotene and zygotene cells were isolated in combination, and were >90% pure. Neither pachytene nor leptotene/zygotene fractions exhibited evidence of any contaminating somatic cells. RNA was extracted from these STA-PUT purified cells using TRIzol (Life Technologies), and was used as the source material for mRNA sequencing, small RNA sequencing, and qRT-PCR. Illumina kits were used for preparation of both RNA-Seq (Illumina TruSeq RNA Seq)and Small RNA-Seq libraries (Illumina TruSeq Small RNA Seq).