Fusarium ear rot (FER), caused by Fusarium verticillioides, is a destructive fungal disease of maize. FER resistance is a complex, quantitatively inherited trait controlled by multiple minor-effect genes. In this study, we employed two recombinant inbred line (RIL) populations with the common resistant parental line CML304 to identify FER-resistance loci. Initial QTL analysis identified 23 FER-resistance QTL, each explaining 5.21%–30.51% of the total phenotypic variation. Notably, one major QTL, qRfv2, on chromosome 2 was repeatedly detected, accounting for 11.92%–30.51% of the total phenotypic variation. qRfv2 was fine mapped to an interval of 1.01 Mb, flanked by the markers IDP8 and IDP10. qRfv2 is a semi-dominant resistance gene that could reduce the disease severity index (DSI) by 12.4%–20%, suggesting its potential for enhancing FER resistance in maize. Transcriptome analysis showed that 22 of the 28 annotated functional genes in the qRfv2 region displayed differential expression between parental lines in response to FER. One of the candidate genes, ZmLOX6, was validated to presumably provide a positive effect on FER resistance. Our study provides a basis for the potential cloning and application of FER resistance genes in maize breeding.