RNA isolation / purification Cells - primary mouse cortical neurons

When extracting nucleic acids from cell cultures, thorough homogenization of cells via vortexing in lysis buffer is very necessary. Choose the best RNA isolation method keeping in mind the downstream applications, generally, column-based isolations result in clean and concentrated RNA samples. Downstream applications like sequencing and cDNA synthesis require high-quality RNA, always treat the samples with DNases and check their integrity by running a gel.

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5 years ago

5 years ago by Ralf Friedmann Switzerland

I am having trouble using the QiageN RNAeasy mini kit for CD34+ cells. Any tips?

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Found 4 matching solutions for this experiment

Aurum™ Total RNA Mini Kit

Bio-Rad Laboratories

Upstream tips	Protocol tips	Downstream tips
- This kit is designed to process upto 2x10^6 cells - 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
- This kit is designed to process upto 2x10^6 cells - 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

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

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

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

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