Apple Latent Spherical Virus (ALSV) Vector as a Tool for Reverse Genetic Studies and Non-transgenic Breeding of a Variety of Crops

Apple latent spherical virus (ALSV) has recently emerged as an efficient system for reverse-genetic tool of virus-induced gene silencing (VIGS), gene expression, and potential epigenetic breeding through virus-induced transcriptional gene silencing (VITGS) of crops. ALSV does not necessarily infect all plant species, but it infects to a variety of crops such as apple, pear, cherry, soybean, pea, cucumber, watermelon, petunia, Eustoma, and Japanese gentian with the current protocol, at relatively high infection rates. Virus vectors are routinely constructed in binary plasmids, agro-infiltrated to Nicotiana benthamiana, propagated in Chenopodium quinoa, concentrated, RNA extracted, and introduced into crops by gold particle bombardment (biolistic inoculation). Methods and tips of virus preparation and infection are explained in detail in this chapter. We are recently testing whether VITGS technology is applicable to any gene of any plant species. Unexpected problems observed here are shown, and possible solutions to overcome these problems will be discussed.