YDV Multiplex PCR v1

This protocol describes the process of carrying out a multiplex PCR assay followed by capillary electrophoresis to detect C/BYDV and determine species (BYDV-MAV, BYDV-PAS, BYDV-PAV, CYDV (RPS or RPV)). The starting point for this is cDNA that has been synthesized from nucleic acid extracted from single aphids or leaf material - an internal control targeting GAPDH is included and primers can be selected depending on the sample matrix (aphid or barley).The BYDV primers were taken from: Sõmera, M., Massart, S., Tamisier, L., Sooväli, P., Sathees, K. and Kvarnheden, A., 2021. A survey using high-throughput sequencing suggests that the diversity of cereal and barley yellow dwarf viruses is underestimated.Frontiers in Microbiology,12, p.673218. In-house sequence data was used to modify primers where necessary, add a primer for CYDV and an internal control for both aphid and barley samples. The CYDV primers captures both RPS and RPV (high resolution melt analysis may be required to distinguish these if required).