GSM4943503: rrmd_lv3; Monodelphis domestica; RNA-Seq
Tissue was dissected at indicated ages (see section ”Samples” in GEO metadata). RNA for mouse, rat, rhesus and human was extracted using the RNeasy protocol from QIAGEN; for opossum, a TRIzol/chloroform-based protocol was used. Tissues were stored at -80 C until library preparation. We used 5µg of RNA per tissue as starting material for library preparation, which were treated with RNase R for 1 h at 37 ̊C to degrade linear RNAs, followed by RNA purification with RNA Clean & Concentrator-5 kit (Zymo Research) according to the manufacturer's protocol. Libraries were prepared from the purified RNA with the Illumina TruSeq Stranded Total RNA with Ribo-Zero Gold according to the protocol with the following modifications to select larger fragments: 1). Instead of the recommended 8 min at 68 ̊C for fragmentation, we incubated samples for only 4 min at 68 ̊C to increase the fragment size; 2). In the final PCR clean-up after enrichment of the DNA fragments, we changed the 1:1 ratio of DNA to AMPure XP Beads to a 0.7:1 ratio to select for binding of larger fragments. Libraries were analyzed on the fragment analyzer for their quality and sequenced with the Illumina HiSeq 2500 platform.
GEO Accession
GSM4943503