Executive Summary : | Tomato, the second-highest-consumption crop globally, faces challenges due to biotic and abiotic factors. The Tomato leaf curl virus (ToLCV), a bacterium transmitted by whiteflies, is the most destructive and causes 20-100% yield loss. Despite the significant damage caused by ToLCV, no potential control strategy has been developed, except for controlling whiteflies using insecticides. Various approaches, including integrated pest management, stringent quarantine, traditional breeding, and transgenic technology, have been applied to control ToLCV. ToLCV-resistant linked QTLs (QTLs) have been identified, including Ty-1 to Ty-6 in wild tomatoes. The Ty-5 locus contains the SlPelo gene, which encodes the mRNA surveillance factor called pelota (PELO), which plays a key role in ribosome recycling during protein synthesis. To meet the increasing global population demand, traditional breeding is time-consuming and lacks 100% effectiveness. To address this issue, the engineering of ToLCV resistance tomato via clustered regularly interspaced short palindromic repeats (CRISPR) with Cas9 nuclease (CRISPR/Cas9) is a promising tactic. CRISPR/Cas9 is advanced, cost-effective, more accurate, and effective in multiple site genome editing. In this work, a novel CRISPR-Cas9 tool with multiplexed guide RNA (gRNA) will be used to edit SlPelo at two or three sites simultaneously for ToLCV resistance. This approach is promising for developing resistance against ssDNA/dsDNA viruses in various crops. |