Ion Total RNA-Seq Kit v2

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	100 ng–50 μg total RNA, we recommend using the DynabeadsTM mRNA DIRECTTM Micro Kit Micro Kit (Cat. No. 61021). 50–400 μg total RNA, we recommend performing two rounds of oligo(dT) selection of the poly(A) RNA using the MicroPoly(A)PuristTM Kit (Cat. No. AM1919). Also, confirm the absence of 18S and 28S rRNA; for example, check the profile of the poly(A) RNA on an AgilentTM 2100 BioanalyzerTM instrument.

Protocol tips

100 ng–50 μg total RNA, we recommend using the DynabeadsTM mRNA DIRECTTM Micro Kit Micro Kit (Cat. No. 61021). 50–400 μg total RNA, we recommend performing two rounds of oligo(dT) selection of the poly(A) RNA using the MicroPoly(A)PuristTM Kit (Cat. No. AM1919). Also, confirm the absence of 18S and 28S rRNA; for example, check the profile of the poly(A) RNA on an AgilentTM 2100 BioanalyzerTM instrument.

Publication protocol

Total RNA was extracted from cells using TriPure isolation reagent (Roche, Indianapolis, U.S.A.), and used for RNA-seq and quantitative (q)RT-PCR. For RNA-seq, mRNA was purified by removal of ribosomal RNA using a RiboMinus Eukaryote kit for RNA-seq (ThermoFisher Scientific, Rockford, U.S.A.). RNA-seq libraries were then prepared using an Ion Total RNA-Seq Kit Ver2 (ThermoFisher Scientific). Samples were amplified by emulsion PCR and Ion Sphere particles were concentrated. Then, samples were analyzed on a next generation sequencer (Ion Proton, ThermoFisher Scientific). The obtained sequence data were analyzed using CLC Genomics Workbench software 10.0.1 (QIAGEN, Redwood City, U.S.A.).

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Manufacturer protocol

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