RNA isolation / purification Tissue - Human Subcutaneous

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.

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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?

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Discussion

4 years ago

4 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?

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Found 3 matching solutions for this experiment

Aurum™ Total RNA Mini Kit

Bio-Rad Laboratories

Upstream tips	Protocol tips	Downstream tips
- Before using lysis solution, add 500 ul of β-mercaptoethanol (β-ME) to the solution for a final concentration of 1%.	- To perform efficient elution preheat the elution solution at 70C in water bath priorto the elution step. - If there is high genomic DNA contamination, increase DNAse I digestion time and use only the DNAse dilution solution provided in the kit	- For better downstream application add appropriate volume of 95-100% ethanol to the wash solutions before intial use. - If elute volume is >80ul, there is a possibility of ethanol contamination. To reduce this, add 1-3min of centrifugation time after the final wash step.

Upstream tips
- Before using lysis solution, add 500 ul of β-mercaptoethanol (β-ME) to the solution for a final concentration of 1%.
Protocol tips
- To perform efficient elution preheat the elution solution at 70C in water bath priorto the elution step. - If there is high genomic DNA contamination, increase DNAse I digestion time and use only the DNAse dilution solution provided in the kit
Downstream tips
- For better downstream application add appropriate volume of 95-100% ethanol to the wash solutions before intial use. - If elute volume is >80ul, there is a possibility of ethanol contamination. To reduce this, add 1-3min of centrifugation time after the final wash step.

Manufacturer protocol Publication protocol 1 Relevant paper

High Pure RNA Isolation Kit

Roche Lifesciences

Upstream tips	Protocol tips	Downstream tips
- To decrease RNase activity,process starting material immeditely or store at 80C. - To increase nucleic yield or purity store all buffer at +15C to +25C.	- To decrease RNase activity,use eluted RNA directly in downstream procedure or store at -80C immediately

Upstream tips
- To decrease RNase activity,process starting material immeditely or store at 80C. - To increase nucleic yield or purity store all buffer at +15C to +25C.
Protocol tips
- To decrease RNase activity,use eluted RNA directly in downstream procedure or store at -80C immediately

Manufacturer protocol Publication protocol 1 Relevant paper

PureLink™ RNA Mini Kit

Thermo Fisher Scientific

Upstream tips	Protocol tips	Downstream tips
	- Depending on the material you want to isolate RNA from, please refer to the manual for additional reagents/lab equipment that are required	- DNase is not supplied with this kit, a DNase kit is available which is compatible with this kit

Protocol tips
- Depending on the material you want to isolate RNA from, please refer to the manual for additional reagents/lab equipment that are required
Downstream tips
- DNase is not supplied with this kit, a DNase kit is available which is compatible with this kit

Manufacturer protocol Publication protocol 1 Relevant paper

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