RNeasy Mini Kit

RNA isolation / purification Tissue - Rat ileum

Experiment

RNA isolation / purification Tissue - Rat ileum

Product

RNeasy Mini Kit from Qiagen

Manufacturer

Qiagen

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Tips Publication protocol Reviews Discussion Papers Manufacturer protocol Videos

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

The hypothalamus was quickly dissected from a semi-frozen brain. Total RNA was isolated from the hypothalamus, duodenum, jejunum, and ileum using RNeasy mini-columns (Qiagen, Valencia, CA). Genomic DNA was eliminated by DNase I on column treatment with RNase-free DNase set (Qiagen, Valencia, CA). RNA integrity was confirmed by visualizing an approximate 2/1 ratio of 28S to 18S band on the 1% agarose gel stained by ethidium bromide. RNA purity was confirmed with absorbance ratio of >2.0 at 260nm/280nm using NanoVue (GE Healthcare, Piscataway, NJ). 1 μg of total RNA was used to generate cDNA using iScript following manufacturer's protocol (Bio-Rad, Hercules, CA). cDNA was made from all samples for each tissue at the same time to minimize experimental variations.

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Discussion

4 years ago

Author: Paul G. Macon United States

RNA isolation from tissue

How do I extract RNA from animal tissue without using liquid nitrogen? I tried the RNA extraction by using the TRIzol reagent and I homogenize the tissue using polytron homogenizer at room temperature for 30secs is this correct?

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Discussion

5 years ago

Author: Aaron Stege Netherlands

Problem in phase separation after using serum/plasma kit

I used a serum/plasma kit for my serum samples. After the phase separation the samples should have 3 phases: a colourless aqueous phase, a white interphase and a red organic phase. However, in some of my samples there was no aqueous phase unless I wait for an extended period of time. How can I circumvent this problem?

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Papers

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Manufacturer protocol

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

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