SBIR Phase II: Generating HLB and canker resistant citrus trees using CRISPR/Cas9

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2). The broader impact of this Small Business Innovation Research (SBIR) Phase II project is to help the US citrus industry mitigate the devastating effects of two diseases: Huanglongbing (HLB) and citrus canker, both of which have killed millions of citrus trees in Florida, cost the state billions in lost revenue, and caused the loss of thousands of jobs. HLB has also spread to other states including California, Texas, Arizona, and Georgia. This Phase II project will produce non-GMO, disease-resistant citrus trees using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) precision breeding technology. These new varieties will boost the US citrus industry, especially in Florida where HLB and canker have led to a 79% production decline. The proposed project aims to combine effector-interactome and CRISPR to create a commercially valuable new citrus variety for growers. This project leverages research that identified citrus genes that interact with bacterial effector proteins causing disease development. The project explores disruption of the pathogen-citrus interactions by a multiplex CRISPR system and the associated potential for disease resistance in commercially valuable citrus varieties. This project will generate citrus plants with edits to promising target genes and test those new varieties for resistance and other agronomic traits in greenhouse and field trials. The methods and approaches are multiplex CRISPR plasmid constructions, and citrus protoplast transformation and regeneration to create non-transgenic resistant citrus varieties. The testing protocol calls for grafting to inoculate the new varieties with HLB to confirm resistance and clonal micropropagation for large-scale experiments and field trials. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.