|Powell, Joseph - Larry
|Arias De Ares, Renee
Submitted to: Plant Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/29/2016
Publication Date: 1/5/2017
Citation: Power, I.L., Dang, P.M., Sobolev, V., Orner, V.A., Powell Jr, J.L., Lamb, M.C., Arias De Ares, R.S. 2017. Characterization of small RNA populations in non-transgenic and aflatoxin-reducing-transformed peanut. Plant Science. 257:106-125. doi:10.1016/j.plantsci.2016.12.013.
Interpretive Summary: Aflatoxins that accumulate in crops after infection with the fungus Aspergillus flavus and A. parasiticus are a serious risk for human health and food security. We have used RNA interference (RNAi) technology and engineered peanut plants to make small RNA interference (siRNA) to target genes that those fungi use to make aflatoxins. We also developed a method to test the effectiveness of RNAi controlling aflatoxin in transgenic peanut seeds. The result was overall 80-100 % reduction in aflatoxin accumulation. This is the first report of peanut-plant RNA-mediated control of aflatoxins, and the first example of using RNAi in plants to control mycotoxins produced by plant pathogens.
Technical Abstract: Aflatoxins are powerful carcinogenic secondary metabolites produced mainly by Aspergillus flavus and A. parasiticus. These mycotoxins accumulate in crops and pose a serious risk to food safety and human health. No consistently effective method exists to control aflatoxins in crops. RNA interference (RNAi) is a powerful biotechnology tool; RNAi signals travel within the plant and can silence genes inside pathogens living in close contact with the plant. Key challenges in evaluating the effectiveness of RNAi to control aflatoxins in peanuts are: unpredictability of aflatoxin accumulation, inherently-high variability of RNAi silencing among transgenic events in plants, and the range of seed maturity stages. No method is available to evaluate new peanut breeding lines for a no-aflatoxin accumulation trait other than multi-year field testing and analysis of large number of samples. We developed a method for testing effectiveness of RNAi silencing using few seeds of individual transgenic events. We made RNAi construct targeting five genes in the aflatoxin-biosynthesis pathway, transformed the construct into peanut plants using Agrobacterium, applied A. flavus to transgenic seeds, and quantified aflatoxin accumulation by ultra-performance liquid chromatography. Overall, RNAi peanut lines showed 84-100 % less aflatoxin B1 than the controls, and variable response by seed-maturity stages. The method described here, can be used to screen transgenic events for host-plant RNA-mediated silencing of aflatoxin synthesis. This is the first report of peanut-plant RNA-mediated control of aflatoxins, and the first of its kind on the use of RNAi in plants to control mycotoxins produced by plant pathogens.