To be detailed, 2 μL of pseudotyped viruses containing the SARS-CoV-2 N gene fragment were used template and mixed with qPCR mix containing 10 μL of 2 × T5 Fast qPCR Mix, 0.8 μL of forward primer/reverse primer (10 μM), 0.4 μL of DNA probe (10 μM), 0.4 μL of 50 ...
In brief, 10 µL of reaction mix containing 5 µL of 2× PerfeCTa qPCR ToughMix (Quantabio, USA); 1.5 µL of primer and probe mixture that comprised 400 nM each of the forward and reverse primers and 200 nM each of the fluorescent hydrolysis probes; 3 µL of 1:10 diluted cD...
The RT-qPCR was performed using the SuperScript III Platinum One-Step Quantitative RT-PCR System (Invitrogen) with the US-CDC 2019-nCoV-N1 combined primer and probe mix. A one-step RT-PCR consisting of a 30-min RT step at 50 °C, 5 min of Taq polymerase inhibitor inactivation at 95 ...
qPCR Probe-based qPCR was prepared by mixing the following reagents: 1 µL of cDNA, 1× PrimeTime Gene Expression Master Mix (IDT #1055772), 1× primers/probe mix, and nuclease-free water to a final volume of 10 µL. Three technical replicates were performed for each sample. qPCR ...
Glycosyltransferase mRNA expression levels were quantified by RT-qPCR using TaqManTM Universal PCR Master Mix II, no UNG, and the specific TaqManTM gene expression assays, listed below, in a 7500 Fast Real-Time PCR System (Applied Biosystems, Foster City, CA, USA; Therm...
qPCR Probe-based qPCR was prepared by mixing the following reagents: 1 µL of cDNA, 1× PrimeTime Gene Expression Master Mix (IDT #1055772), 1× primers/probe mix, and nuclease-free water to a final volume of 10 µL. Three technical replicates were performed for each sample. qP...
In case 1, due to previous recurrent bacterial infections and SARS-CoV-2 evidence with a high CT value, there was probably a persistent infection with SARS-CoV-2 in an immunocompromised patient. In a previous hospitalization, four to six weeks before death, the patient tested several times pos...
(amplicon of 124 bp) showed lower transcript intensity in the two patients for both amplifications, particularly in the daughter compared with two controls (Figure 4A). We quantified theFGFR2transcript in the two patients and healthy controls using relative qPCR (Figure 4B). Using a probe that...
Briefly mix the first tube and prepare a 1:100 dilution by transferring 200 μL of the first to the second tube. Then transfer 200 μL of the 1:100 dilution to the next tube in the series. Repeat the procedure to make a serial 1:10 dilution of the lung suspension, e.g., from ...
Three controls were included in the qPCR assay: a no-template control (nuclease-free H2O), a negative control (human WT control DNA, reference 4451855, Ambion, Austin, TX, USA), and a positive control (included in the kit). Good laboratory practices were maintained to limit contamination, ...