RNA was purified using the NucleoSpin RNA II Kit or Competitor Q’s RNeasy Mini kit. 10 µl of purified RNA was treated with RNase A and loaded onto a 1% TAE agarose gel to reveal any contaminating genomic DNA. Genomic DNA was completely eliminated from samples purified with the NucleoSpin RNA II kit, but RNA purified with the RNeasy Mini kit showed evidence of genomic DNA contamination.
RNA was purified using the NucleoSpin RNA II Kit or Competitor Q’s RNeasy Mini kit. 10 µl of purified RNA was treated with RNase A and loaded onto a 1% TAE agarose gel to reveal any contaminating genomic DNA. Genomic DNA was completely eliminated from samples purified with the NucleoSpin RNA II kit, but RNA purified with the RNeasy Mini kit showed evidence of genomic DNA contamination.
How Does NucleoSpin RNA II Remove Genomic DNA Contamination?
If you want to isolate high-quality RNA, it is important to prevent degradation of the RNA and to eliminate genomic DNA. Unlike Competitor Q’s RNeasy Kits, NucleoSpin RNA II includes the following:
- An optimized DNase I treatment to eliminate genomic DNA contamination
- Filters/shredders to homogenize samples and reduce viscosity
Without these components, RNA purified with Competitor Q’s RNeasy kits exhibits significant DNA contamination.
Total RNA Purification from Just a Few Cells
With NucleoSpin RNA II, you can detect transcripts from very small numbers of cells—in this example, RNA was isolated from as few as 10 HeLa cells.
Total RNA was isolated from the indicated number of HeLa cells with NucleoSpin RNA II (elution volume: 40 µl). RT-PCR reactions were carried out with 3 µl of total RNA and beta-actin primers. The RT-PCR product was 626 bp.
Simple Protocol for DNA Removal
NucleoSpin RNA II uses a simple spin-column procedure that is probably already familiar to you. First, cells are lysed in a buffer that inactivates RNases (which are present in virtually any biological material) and creates appropriate conditions for RNA to bind to the silica membrane. After lysis, homogenization and reduction of viscosity are achieved by filtration with NucleoSpin Filter units (shredders), which are provided with the kit. Residual genomic DNA is removed by on-column digestion with rDNase, which is also included in the kit.
What is NucleoSpin RNA II?
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Technology
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Silica membrane
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Format
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Mini spin columns
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Starting material
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5 x 106 cultured cells or 109 bacterial cells or 30 mg tissue
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Fragment size
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>200 bases
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Yield
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14–70 µg total RNA
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Elution volume
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40–100 µl
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Time
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30 min/6 preps
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