Begomoviruses are exclusively transmitted by the whitefly Bemisia tabaci in a persistent circulative manner and pose serious threat to the production of important cash crops worldwide. Here, we describe the protocols to study how the whiteflies transmit begomoviruses, including maintenance of whitefly and species identification, preparation of tomato plants infected with virus, acquisition and transmission of virus by whiteflies, dissection of whitefly organs, analysis of virus quantity in whiteflies, immunofluorescence and fluorescence in situ hybridization (FISH) detection of virus in whitefly tissues, membrane feeding of double-stranded RNA (dsRNA) or antibody and microinjection of dsRNA in whiteflies.