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1 year ago
1 year ago by Ralf Friedmann
I am having trouble using the QiageN RNAeasy mini kit for CD34+ cells. Any tips?
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|The total RNA was extracted from cultured cells after 24 h incubation with various concentrations of IL-6 or IL-6/IL-8 using a High Pure RNA Isolation kit (Roche, Basel, Switzerland), according to manufacture's protocol. The extracted RNA was purified and diluted in DNase and RNase-free water. The quality and quantity of isolated RNA was measured using a spectrophotometer NanoDrop® (Thermo Fisher Scientific, Inc.).|
|Total RNA was isolated from A2780 and A2780CP20 ovarian cancer cell lines with the GenElute Mammalian Total RNA Purification Kit (Sigma-Aldrich). The RNA concentration was read in a nanodrop. RNA quality was verified in a Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA). One hundred nanograms of total RNA were used to cDNA synthesis.|
|- To yield hign 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.