mirVana™ miRNA Isolation Kit, with phenol

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	- For RNA to dissolve better and to yeild high levels, preheat elution buffer at 95C. - Addition of β-mercapto ethanol in lysis buffer can be skipped because the phenol will do the job of nuclease inactivation. - To capture more amount of mRNA, modify the amount of lysis buffer:ethanol (from 1:1 to 1:1.5) during the binding step.	- Do vortex for 2-plus minutes during Acid-Phenol:Chloroform extraction step. This will facilitate the removal of tightly bound proteins typically associated with RNA.

Protocol tips
- For RNA to dissolve better and to yeild high levels, preheat elution buffer at 95C. - Addition of β-mercapto ethanol in lysis buffer can be skipped because the phenol will do the job of nuclease inactivation. - To capture more amount of mRNA, modify the amount of lysis buffer:ethanol (from 1:1 to 1:1.5) during the binding step.
Downstream tips
- Do vortex for 2-plus minutes during Acid-Phenol:Chloroform extraction step. This will facilitate the removal of tightly bound proteins typically associated with RNA.

Publication protocol

Total RNA from the samples was isolated by using either Ambion’s miRVana kit (AM1560, Life Technologies) or Trizol (15596-018, Life Technologies, USA). DNA contamination in the total RNA was removed by DNase I (AMPD1, Sigma-Aldrich, USA) treatment. Complementary DNAs (cDNA) from the RNA samples were prepared by using Superscript cDNA synthesis kit (18080, Life Technologies, USA). High throughput miRNA cDNAs were prepared by using NCode™ miRNA First-Strand cDNA (MIRC-50, Life Technologies, USA) synthesis kit.

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Manufacturer protocol

Download the product protocol from Thermo Fisher Scientific for mirVana™ miRNA Isolation Kit, with phenol below.

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