Improved method for assaying maize plant resistance to maize rough dwarf disease by artificial inoculation and real-time RT-PCR

The study of maize rough dwarf disease (MRDD) and breeding for resistance requires inoculation of maize plants by means of planthoppers. The plant age, insect density and inoculation duration are main factors in the success of maize rough dwarf disease inoculation. These parameters were tested using a susceptible maize inbred line Ye478. Using one or two-leaf plants, 15 planthoppers per plant and a five day inoculation duration, the line Ye478 was the most susceptible with 100% diseased plants; F112132 was moderately susceptible with 60% diseased plants and 90110 and F022411 were resistant without any disease. The results were consistent with those from six years of field studies. Using enzyme-linked immunosorbent assay (ELISA) and real-time quantitative RT-PCR, rice black-streaked dwarf virus was detected in severely diseased plants. The plants were rated from 0 to 3 according to their symptoms at the time of flowering. Plants scoring 0, 1 and 2 could not be distinguished by ELISA, only by real-time quantitative RT-PCR. All of the plants with a score of 3 were positive by ELISA and real-time quantitative RT-PCR. The significant differences in the average viral contents in plants with different symptom ratings could be distinguished by using real-time RT-PCR.