Reliable detection and quantification of barley and cereal yellow dwarf viruses (YDVs) is a critical component in managing yellow dwarf diseases in small grain cereal crops. The method currently used is enzyme-linked immunosorbent assay (ELISA), using antisera against the coat proteins that are specific for each of the various YDVs. Recently, quantitative real-time reverse-transcription polymerase chain reaction (Q-RT-PCR) has been used to detect bacterial and viral pathogens and to study gene expression. We applied this technique to detect and quantify YDVs using primers specific for Barley yellow dwarf virus-PAV (BYDV-PAV) and Cereal yellow dwarf virus-RPV (CYDV-RPV) coat protein genes because of the higher sensitivity of RT-PCR and the advantage of using a real-time PCR instrument. This Q-RT-PCR was used to detect BYDV and CYDV, and to examine disease development in a resistant wheatgrass, a resistant wheat line, a susceptible wheat line, and a susceptible oat line. BYDV-PAV and CYDV-RPV were detected as early as 2 and 6 h, respectively, in susceptible oat compared with detection by ELISA at 4 and 10 days postinoculation. BYDV-PAV RNA accumulated more rapidly and to a higher level than CYDV-RPV RNA in both oat and wheat, which may account for PAV being more prevalent and causing more severe viral disease than CYDV. Q-RT-PCR is reproducible, sensitive, and has the potential to be used for examining yellow dwarf disease and as a rapid diagnostic tool for YDVs.

Stable introgression of agronomically important traits into crop plants through wide crossing often requires the generation and identification of translocation lines. However, the low efficiency of identifying lines containing translocations is a significant limitation in utilizing valuable alien chromatin-derived traits. Selection of putative wheatgrass-wheat translocation lines based on segregation ratios of progeny from gamma-irradiated seed using a standard phenotypic analysis resulted in a low 4% success rate of identifying barley yellow dwarf virus (BYDV) resistant and susceptible translocation lines. However, 58% of the susceptible progeny of this irradiated seed contained a Thinopyrum intermedium chromosome-specific repetitive sequence, which indicated that gamma-irradiation-induced translocations occurred at high rate. Restriction fragment length polymorphism (RFLP) analysis of susceptible lines containing alien chromatin, their resistant sister lines and other resistant lines showed that more than one third of the progeny of gamma-irradiated double monosomic seeds contained wheatgrass-wheat translocations. Genomic in situ hybridization (GISH) analysis of selected lines confirmed that these were wheatgrass-wheat translocation lines. This approach of initially identifying BYDV susceptible deletion lines using an alien chromosome-specific repetitive sequence followed by RFLP analysis of their resistant sister lines efficiently identified resistant translocation lines and localized the BYDV resistance to the distal end of the introgressed Th. intermedium chromosome.