In December 2019, an increase in the number of severe pneumonia cases was reported in China, more specifically in the Hubei Province. Both of the proposed assays are less sensitive than the current gold standard however, our data shows a low sensibility variation, suggesting that these methods could be used by laboratories as a lower cost molecular method for SARS-CoV-2 diagnosis. The average Ct variation between SYBR Green and TaqMan was 1.92 and the highest Ct detected by conventional PCR was 35.98.

The conventional PCR detected until 10 7 dilution, both assays detected the majority of the 63 samples, 98.42% of positivity in SYBR Green, and 93% in conventional PCR. The SYBR Green-based assay was able to detect all 8 dilutions of the isolate. The SYBR Green assays and conventional RT-PCR have not shown amplification of the 17 respiratory samples positives for other viruses. The cross-reactivity test was performed with 17 positive samples for other respiratory viruses, and the sensibility test was performed with 8 dilutions (10 based) of SARS-CoV-2 isolated and 63 SARS-CoV-2-positive samples. (2020) were adapted in SYBR Green assay and RT-PCR conventional. E gene primers previously determined for TaqMan assays by Colman et al. The objective of this study was to standardize lower cost molecular methods for SARS-CoV-2 identification. However, it is a high-cost assay, inaccessible for the majority of laboratories around the world, making it difficult to diagnose on a large scale. TaqMan-based real-time RT-qPCR is currently the gold standard for COVID-19 diagnosis. In March 2020, WHO declared a pandemic state due to SARS-CoV-2 having spread. Daniella Gregorio Bonfim Prado Silva 3,.Tighter reproducibility allows for greater significance in analyzing low-abundant transcripts and smaller fold changes.Lower cost alternatives for molecular diagnosis of COVID-19: conventional RT-PCR and SYBR Green-based RT-qPCR However, PowerUp SYBR Green Master Mix, with its dual hot-start Taq DNA polymerase, demonstrates excellent reproducibility over a wide dynamic range for a variety of targets tested (Figure 2). Reproducibility is another important measure of data quality in real-time PCR reactions, and reproducibility often decays at low template concentration where the impacts of variability are exacerbated. The high specificity of the PowerUp SYBR Green Master Mix allows you to spend less time and money optimizing and redesigning primers to get high-quality data. Validation of specificity in SYBR reactions is essential to data quality and validity (Bustin et al 2009). In contrast, nonspecific amplification was seen for some of the same targets in several competitorsвЂ™ mixes as shown by multiple peaks in the melt curve (Figure 1). In an evaluation of 24 different primer sets with PowerUp SYBR Green Master Mix, a single melt curve was obtained in 100% of reactions without the need for primer optimization or redesign. Formulation of the Dual-Lock Taq into a convenient 2X pre-mix containing an optimized buffer system, proprietary additives, and detergents further improves the specificity of PowerUp SYBR Green Master Mix. Providing tight control over Taq activation, this dual hot-start approach helps prevent undesirable early activity of the polymerase at low temperatures that can lead to non-specific amplification. PowerUp SYBR Green Master Mix contains proprietary Dual-Lock Taq DNA Polymerase enzyme, which uses a combination of two hot-start mechanisms to control its activity.

Powerup sybr green master mix