<p>The whitefly-transmitted begomovirus can devastate <i>Solanaceae</i> crops worldwide. Despite a strong demand for the genetic introgression of begomovirus resistance, only the begomovirus resistance gene in tomatoes and peppers has been cloned. Here, we aimed to identify a begomovirus resistance gene in eggplant (<i>Solanum melongena</i>). Previously, we identified accession No.820 as a resistance source against tomato yellow leaf curl Kanchanaburi virus (TYLCKaV). A dominant locus, <i>Eggplant yellow leaf curl disease virus resistance 1</i> (<i>Ey-1</i>), conferring resistance against TYLCKaV was identified on chromosome 1 by genetic mapping using the F₂ and F_2:3 segregating populations obtained from cross-pollination of No.820 and begomovirus susceptible No.47. From whole-genome and transcriptome sequencing of No.820 and No.47, we selected 5 genes as candidates among 10 genes on the final 113-kb target region. Reverse genetic analysis using virus-induced gene silencing (VIGS) of these five candidate genes in No.820 revealed that silencing of <i>SmNEN3,</i> which encodes a DEDDh family exonuclease protein, resulted in loss of resistance. Comparison of the genomic and transcript sequences of <i>SmNEN3</i> from No.820 and No.47 revealed a single amino acid deletion and nonsynonymous mutations that most likely contribute to begomovirus resistance. No.820 is a highly valuable genetic resource with dominant resistance to begomovirus, and the new DNA markers will greatly aid marker-assisted breeding.</p>