Rna clean concentrator 5 column kits

Cells were lysed in Qiazol (QIAGEN) and total RNA was purified using the miRNEasy mini kit (QIAGEN) according to the manufacturer’s instructions. Tissues were lysed in Qiazol (QIAGEN) and total RNA was purified using Tissuelyser II (QIAGEN), as per the manufacturer’s instructions. Small RNAs (<200nt) size-fractionated using RNA Clean & Concentrator 5 column kits (Zymo), as per the manufacturer’s instructions. Testes of 8 weeks old mice were grinded under liquid nitrogen and RNA was extracted using Trizol (Thermo Fisher Scientific) according to the manufacturer’s instructions but using 80% EtOH during the wash step.

Cells were lysed in Qiazol (QIAGEN) and total RNA was purified using the miRNEasy mini kit (QIAGEN) according to the manufacturer’s instructions. For mRNA detection, 1 μg of purified total RNA was reverse transcribed using the high-capacity cDNA reverse transcription kit (Applied Biosystems). For specific miRNA quantification, we size-fractionated small RNAs (< 200nt, containing pre-miRNAs) using RNA Clean & Concentrator 5 column kits (Zymo), as per the manufacturer’s instructions. miRNAs were reverse transcribed with miScript II RT kit (QIAGEN). Primers were designed to anneal on the 5p or 3p arm of the miRNA of interest (Supplementary Table S1). miRNAs were quantified using Fast SybrGreen PCR mastermix (Applied Biosystems) according to the manufacturer’s instructions.