RNA isolation / purification Tissue - Mouse Muscle

Isolating RNA from tissues and paraffin-embedded tissue samples can be challenging due to cross-linking of biomolecules and fragmented nucleic acids. The best solution is to slice the tissues into smaller pieces and make a homogenate solution (using tissue homogenizer or grinding liquid nitrogen frozen samples) in presence of RNAse inhibitors. The homogenization process should be carried out on dry ice to maintain the integrity of RNA.

7 Matching solutions
2 Discussions

Found 2 discussions for this experiment

Discussion

4 years ago

4 years ago by 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?

Comment View 2 comments

Discussion

5 years ago

5 years ago by 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?

Comment View 2 comments

Share your thoughts or question with experts in your field by adding a discussion!

Found 7 matching solutions for this experiment

RNeasy Plus Mini Kit

Qiagen

Upstream tips	Protocol tips	Downstream tips
	- On-column digestion is preferable - For high gDNA elimination spin at 6000rpm for 5 min instead of the quick high speed spin	- 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
- On-column digestion is preferable - For high gDNA elimination spin at 6000rpm for 5 min instead of the quick high speed spin
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.

Manufacturer protocol Publication protocol 1 Relevant paper

RNeasy Mini Kit

Qiagen

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.

Manufacturer protocol Publication protocol 1 Relevant paper

NucleoSpin® RNA

Macherey Nagel

Upstream tips	Protocol tips	Downstream tips
- Aliquot rDNase and store at -20 °C.	- Dry the columns very well after the ethanol wash by adding an additional 2' >10000 rpm centrifuge with no buffer.

Upstream tips
- Aliquot rDNase and store at -20 °C.
Protocol tips
- Dry the columns very well after the ethanol wash by adding an additional 2' >10000 rpm centrifuge with no buffer.

Publication protocol 1 Relevant paper

TriPure Isolation Reagent

Sigma-Aldrich

Upstream tips	Protocol tips	Downstream tips
- Process tissues immediately and freeze at -60c or below for Low RNA degradation	- Adding too little reagent can lead to contamination with DNA

Upstream tips
- Process tissues immediately and freeze at -60c or below for Low RNA degradation
Protocol tips
- Adding too little reagent can lead to contamination with DNA

Manufacturer protocol Publication protocol 1 Relevant paper

miRNeasy Mini kit

Qiagen

Upstream tips	Protocol tips	Downstream tips
	- To capture a higher amount of mRNA, modify the amount of lysis buffer :ethanol (from 1:1 to 1:1.5) during the binding step.

Protocol tips
- To capture a higher amount of mRNA, modify the amount of lysis buffer :ethanol (from 1:1 to 1:1.5) during the binding step.

Manufacturer protocol Publication protocol 1 Relevant paper

TRIzol Reagent

Thermo Fisher Scientific

Upstream tips	Protocol tips	Downstream tips
	- For low 260/230 readings the best approach is to try washing sample (precipitation) with ethanol to desalt it.

Protocol tips
- For low 260/230 readings the best approach is to try washing sample (precipitation) with ethanol to desalt it.

Manufacturer protocol Publication protocol 1 Relevant paper

SV Total RNA Isolation System

Promega

Upstream tips	Protocol tips	Downstream tips
	- Do not freeze-thaw the DNase more than three times after rehydration

Protocol tips
- Do not freeze-thaw the DNase more than three times after rehydration

Manufacturer protocol Publication protocol 1 Relevant paper

Can't find the product you've used to perform this experiment? It would be great if you can help us by Adding a product!

Outsource your experiment

Fill out your contact details and receive price quotes in your Inbox

Outsource experiment