SMN1 and SMN2 copy numbers were determined using TaqMan® droplet digital PCR (ddPCR) on a QX200 system (Bio-Rad Laboratories, Hercules, CA, USA) and ddPCR™ Supermix for Probes (No dUTP) (Cat. No. 1863024; Bio-Rad Laboratories). BCKDHA was used as a reference gene [30]. Primer...
Biodistribution of AAV-syn-GFP DNA was quantified as GFP gene copy numbers per cell using droplet digital PCR (ddPCR) in the liver, kidney, muscle, heart, spleen and lung. Three AAV serotypes were compared after systemic injection of 3 × 109vector genomes per gram (VG/g) in C57BL/...
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Digital PCR was done on a QX200 instrument (Bio-Rad) using the One-Step RT-ddPCR Advanced Kit for Probes (Bio-Rad #1864022) according to the manufacturer’s instructions. Briefly, 22 µl pre-reactions were prepared consisting of 5 µl 4 × supermix, 2 µl reverse transcriptase...
Multiplexed probe-based RT-ddPCR assays were performed using the One-Step RT-ddPCR Advanced Kit for Probes (Bio-Rad, Hercules, CA, USA) and manufacturer instructions for the quantification of SARS-CoV-2 and SARS-CoV-2 variants. The N1/N2 assay targeted the SARS-CoV-2 viral nucleocapsid (N...
RNase-freewater.EachreactionmixwasanalyzedwiththeQXONEDropletDigitalPCR (ddPCR)System(Bio-Rad).Thethermalcyclingconditionswere:45°Cfor10min(reverse transcription);95°Cfor5min;and40cyclesof95°Cfor15sand58°Cfor30s.Theaverage 555 concentrationsofORF1ab,Ngene,andEgenewere4.16×10,5.33×10,and5.04×...
Table S1. Quantitative mass spectrometry data for subgenome-resolved SARS-CoV-2 RNA interactomes, related to Figure 1 Table S2. Analysis of subgenome-resolved SARS-CoV-2 RNA interactomes, related to Figure 1 Table S3. Analysis of strand preference for SND1 (eCLIP) and NSP9 (cRIP) peaks on...
We measured the concentrations of both human (RPP30) cDNA and SARS-CoV-2 cDNA in the contrived samples by performing multiplexed droplet digital PCR (ddPCR) using ddPCR Supermix for Probes (no dUTP) (Bio-Rad, Hercules, CA), the CDC nCoV-19 N1 assay (IDT, Newark, NJ), and a commercia...
form 8, 24, 48, and 72 hpi respectively, were amplified in a final volume of 20 μL ddPCR reaction, in the presence of ddPCR Supermix for Probes (no dUTP) 2X (Bio-Rad, Hercules, CA) and TaqMan 2019nCoV assay kit v1 RealTime-PCR (ThermoFisher, Waltham, MA) for viral Spike seq...
(Promega Corp., Madison, USA)19. The ddPCR master mix consisted of (per sample) 2.5 μL One-Step RT–ddPCR reverse transcriptase, 6.25 μL One-Step RT–ddPCR Supermix, 1 μL 300 mmol/L dithiothreitol, 1 μL of each forward and reverse primers, 0.5 μL probe (20 μM primer...