RNeasy Mini Kit

RNA isolation / purification Tissue - Human Kidney

Experiment
RNA isolation / purification Tissue - Human Kidney
Product
RNeasy Mini Kit from Qiagen
Manufacturer
Qiagen

Protocol tips

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

For the cell lines, RNA was extracted from the cells using the RNEasy® kit (Qiagen), generating high‐quality total RNA (i.e., RIN > 8) that was used as input material for library construction with Illumina TruSeq Stranded mRNA reagents. The samples were sequenced on the Illumina HiSeq2500 platform to a depth of ~20 million reads. Raw sequences were mapped to the human reference genome GrCh38 and further quantified using the Kallisto software (Bray et al, 2016). TPM values for genes were generated by summing up TPM values for the corresponding transcripts generated by Kallisto. All cell line data are available at http://www.ncbi.nlm.nih.gov/bioproject/PRJNA183192.

Procedures for extraction of RNA from tissues, library preparation, and sequencing have been described elsewhere (Uhlén et al, 2015). Briefly, reads were mapped to the human reference genome assembly GRCh38 and quantified using Kallisto version 0.42.4. Normalized expression levels (TPM values) on gene level were obtained by summing the estimated values from the constituent transcripts of each gene, respectively. All tissue data are available at http://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-1733/.

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Discussion

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?

Discussion

4 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.

Download PDF Download manufacturer protocol

Videos

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