RNA-Seq of Miwi[RK/RK]; replicate 2
Total RNA was rRNA-depleted and fragmented for 25 min at 94 C in 2X T4 PNK reaction buffer (140 mM Tris-HCl pH 7.6, 20 mM MgCl2, 10 mM DTT, NEB). The fragmented RNA was further processed into libraries using the SMARTer smRNA kit for Illumina (Takara). The RNA fragments were poly-adenylated and reverse-transcribed with an oligo(dT) primer and a template-switching oligo. Libraries were amplified by 12 PCR cycles.