Figure 2. Stability of three house keeping genes over a 2 month period. GAPDH, Human 18S rRNA and beta actin genes were monitored from saliva samples stored at ambient temperature in Norgen's Saliva RNA Preservative. RNA was isolated from 250 µL of preserved samples on day 0, day 4, day 6, day 10, day 16, day 30 and day 60 using Norgen's Total RNA Purification Kit (Cat. 17200). Next, 4 µL of elution was used in 20 µL of the real-time PCR reaction (SYBR Green I) using CFX96 Touch^TM Real-Time PCR Detection System (Bio-Rad). All of the genes were preserved and stable over 2 month period.

Figure 3. Norgen’s Saliva RNA Preservative is able to successfully preserve saliva RNA samples when used outside of the ideal range of 1:1 at room temperature for 3 days.  As the amount of saliva added to 2 mL of Saliva RNA Preservative increased from 0.2 mL to 3 mL, RNase protein and β-actin gene were monitored for RNA stability using one-step RT-PCR. The results indicated that the variation of the saliva input volumes did not significantly influence the RNA gene detection.

Figure 4. Detection of Viral, Bacterial and Human RNA from Preserved Saliva Samples.  Saliva (2 mL) collected from a single healthy donor was mixed with 2 mL of Saliva RNA Preservative. Norgen’s COVID-19 positive control from the 2019-nCoV TaqMan RT-PCR Kit (E/RdRP gene) (Cat. TM67200) was spiked into the preserved saliva along with HeLa cells (1.6 m cells /ml ) to generate a contrived sample. From the contrived sample, total RNA was purified using Norgen’s Total RNA Purification Kit (Cat. 17200) and 5 genes (16s rRNA, COVID-19 E gene, RdRP-P1 and P2 gene and RP) were analyzed by PCR or RT-PCR to detect viral, bacterial and human genes.