RNeasy Mini Kit

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	- To yield high RNA, give the column an extra wash with both RW1 and RPE buffers (3 total washes with each buffer).	- Include DNAse treatment for 15-20min. - Ensure EtOH is completely evaporated off of the column prior to elution. Adjust time from 1min to 5 min at 60`C. - Use water to elute the RNA that is warmed to ~60`C.

Protocol tips
- To yield high RNA, give the column an extra wash with both RW1 and RPE buffers (3 total washes with each buffer).
Downstream tips
- Include DNAse treatment for 15-20min. - Ensure EtOH is completely evaporated off of the column prior to elution. Adjust time from 1min to 5 min at 60`C. - Use water to elute the RNA that is warmed to ~60`C.

Publication protocol

Total RNA was isolated using the RNeasy Mini Kit (Qiagen) and DNaseI treated (Roche) according to manufacturer’s protocol. The High-Capacity cDNA reverse transcription kit (Applied Biosystems) was used to make cDNA, and PCR analysis was performed using the following primer sets: Vangl1 (5′-GGACTCAAGCCACAACGAGTTGTAT-3′) and (5′-ACTACGAGGCTGAAGTCCAAGC-3′), and Vangl2 (5′-ATGAGCGGGATGACAACTGG-3′ and 5′-ACCTTGAGCGTGAACTGAGG-3′). Quantitative real time PCR was performed in a Bio-Rad iCycler CFX-96 real-time PCR machine using SsoAdvanced Universal Probes Supermix (BioRad) and TaqMan gene-specific primer/probe sets (Applied Biosystems). Levels for VANGL1 (Hs01572998_m1), WNT5A (Hs00998537_m1), and NRDP1/RNF41 (Hs00195064_m1) were normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 4352934E) levels for each sample

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Manufacturer protocol

Download the product protocol from Qiagen for RNeasy Mini Kit below.

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