What methods do you recommend for RNA isolation?
We recommend a column-based method, including:
- Norgen Biotek Total RNA, Soil RNA, or Stool RNA Purification Kits
- Zymo Research QuickRNA™ Kit, ZR Fungal/Bacterial RNA MicroPrep™, or ZR Soil/Fecal RNA MicroPrep™
- Arcturus PicoPure® RNA Isolation Kit
- Ambion PureLink® RNA Mini Kit
- Qiagen RNeasy Kits
Organic methods such as TRIzol® Reagent should be subsequently followed with a column-based clean-up method.
Can I use TRIzol® or other phenol-chloroform based extractions for RNA isolation?
We do not recommend the use of TRIzol or similar methods as any carryover of organic solvents may inhibit downstream enzyme activity. If using TRIzol extracted RNA, we recommend using a column-based purification of the RNA prior to input into the kit.
Can I use carrier RNA during RNA isolation?
We do not recommend the use of carriers during RNA isolation. If a carrier is required, please contact Tecan NGS Technical Support for more information.
Do I need to use high-quality total RNA?
No. The Ovation FFPE WTA System employs a whole transcriptome amplification approach and was designed specifically for use with highly degraded, lower quality RNA samples.
Can I use RNA from sources other than FFPE?
Yes. While the Ovation FFPE WTA System was developed specifically for use with total RNA from FFPE samples, it is possible to use intact, non-FFPE RNA samples as well. Performance with intact total RNA samples below 2 ng may vary.
How much FFPE derived total RNA do I need for amplification?
We recommend total FFPE RNA inputs in the range of 50 to 100 ng. Input amounts outside this range may produce unsatisfactory variable results, especially for more degraded RNA.
Can DNA be used as input for the Ovation FFPE WTA System?
No. The Ovation FFPE WTA System is designed to amplify RNA, not DNA.
Can contaminating genomic DNA interfere with the amplification performance?
Yes. This system is designed to amplify RNA, but contaminating genomic DNA may amplify during the process. For this reason we recommend DNase treatment during RNA purification.
Can I use the Ovation FFPE WTA System on bacterial RNA samples?
The Ribo-SPIA amplification process may work with some bacterial RNAs. However, the kit has not been optimized or validated for this purpose.