Input Recommendations
The protocol is optimized for the following input sample types:
| • | 10–100 ng of purified total RNA |
| • | 20–100 ng RNA input from degraded or FFPE samples (DV200 ≥ 36.5) |
Lower input amounts and lesser quality can reduce library yield.
Include a DNase treatment with the RNA isolation method. The DNase treatment ensures sample purity and accurate quantification. Before starting the protocol, quantify the total RNA using standard methods and assess quality using a fragment analysis method.
For infectious disease and microbiology panels including Respiratory Virus Enrichment Kit (RVEK), and Respiratory Pathogen ID/AMR Panel (RPIP), and Viral Surveillance Panel (VSP):
| • | The panels include both DNA and RNA targets and use an input of total nucleic acid or an equal pool by volume of separately eluted DNA and RNA from the same sample. Do not perform DNase steps as part of the protocol if DNA is included as part of the input (total nucleic acid or mix of separately extracted RNA and DNA). |
| • | Achieving the recommended input of 10–100 ng of total nucleic acid is unlikely from low biomass samples. In this case, a standard volume of 7.5 µl nucleic acid eluate (total nucleic acid or mix of separately extracted RNA and DNA) is recommended. |
