Detection of WT and DVG RNA was performed by RT-qPCR, using WT and DVG specific primers and probes. For WT RNA, the probe and primers were designed to bind to the deleted region in the DVG. DVG-specific primers were designed such that a 113 nt region containing the deleted part of the genome is amplified by primers binding to the Pr gene and the NS1 gene. Amplification of this region in WT virus RNA would result in a significantly larger product (2783 nt). In addition, the DVG recognizing probe was designed such that it would bind to the breakpoint of the deletion. The primer sequences are shown in Supplementary Table 3. DVG or WT RT-qPCR was carried out using the TaqMan RNA-to-Ct One-step RT-PCR kit (Applied Biosystems) or the Luna Universal One-Step RT-qPCR kit (Abcam) on a Step-One-Plus Real-Time PCR thermocycler (Applied Biosystems). The number of RNA genomes was derived from a standard curve produced in each RT-qPCR run using in vitro generated DVG or WT RNA.

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