Improved efficiency of Zea mays agroinoculation with Maize streak virus

Agroinoculation is a technique permitting the transmission of geminivirus g enomes cloned in Agrobacterium tumefaciens into a wide variety of mono- and dicotyledonous host plants. Most geminiviruses are obligately transmitted by insect vector species under natural conditions; therefore, agroinoculati on has greatly simplified the study of this group of viruses. In many cases , agroinoculation has replaced insect transmission, and has been used to co mpare virulence characteristics among viruses. Here we report on the discov ery that, in agroinfectious Maize streak virus constructs, the orientation of cloned viral genomes relative to the Cauliflower mosaic virus 35S (CaMV3 5S) promoter of the binary cloning vector pBI121 can significantly affect a groinfectivity of the constructs. Rates at which plants became symptomatic were significantly higher when agroinoculating maize seedlings with constru cts containing the CaMV35S promoter upstream of the viral replication-assoc iated protein (Rep) gene than when the same viruses were cloned either in t he opposite orientation or into a vector without a strong eukaryotic promot er sequence. Plants infected using the construct with Rep cloned downstream of the CaMV35S promoter also displayed more stunting and, in the early sta ges of the infection, more severe chlorotic streak symptoms.