GSM4160722: Degradome.mouse.testis.wt.adult.Puromycin.rep1; Mus musculus; OTHER
Total RNAs were extracted using mirVana miRNA Isolation Kit (ThermoFisher, AM1560), and rRNAs were depleted from total RNAs with complementary DNA oligomers (IDT) and RNaseH (Invitrogen, Waltham, MA, USA). Degradome: The RNAs were first oxidized at room temperature for 30 minutes with sodium periodate (Sigma, St. Louis, MO, USA) to block the 3´-ends from ligation, and were then size-selected to isolate RNA ≥ 200 nt (DNA Clean & Concentrator™-5, Zymo Research). 5´ adaptors were attached using T4 RNA ligase (Ambion) at 20°C for 3 hours. The ligated products were subjected to rRNA depletion with complementary DNA oligomers (IDT) and RNaseH (invitrogen). The rRNA depleted ligation products were reverse transcribed using a degenerate primer (5´-GCACCCGAGAATTCCANNNNNNNNC-3´). cDNA was amplified by PCR using KAPA HIFI Hotstart polymerase (Kapa Biosystems, Wilmington, MA, USA), and 250–350 nt dsDNA was isolated on 8% native PAGE gels.
GEO Accession
GSM4160722