Development and Application of an SNP Marker for High-Throughput Detection and Utilization of the badh2 Gene in Rice Breeding

<b>Background:</b> As a key rice breeding resource, aromatic rice is widely cultivated in agriculture due to its unique aroma. <i>Badh2</i> mutations cause function loss, enabling rice's characteristic aroma. <b>Methods</b>: In this study, we analyzed several <i>badh2</i> mutation types across 8 <i>japonica</i> and 16 <i>indica</i> aromatic rice lines. Based on the 7 bp deletion in <i>badh2-E2</i> identified in <i>japonica</i> aromatic lines, we developed a multiplex-ready PCR assay for <i>badh2</i> genotyping. Additionally, leveraging the deletion mutation in <i>badh2-E7</i> from the <i>indica</i> aromatic line Yexiang, we designed a KASP marker. <b>Results</b>: All 8 <i>japonica</i> aromatic lines carried a 7 bp deletion in <i>badh2-E2</i>, while 12 <i>indica</i> aromatic lines harbored an 8 bp deletion in <i>badh2-E7</i>, and 4 additional <i>indica</i> aromatic lines exhibited an 8 bp deletion in <i>badh2-E2</i>. The multiplex-ready PCR assay was used to screen 200 individual plants from the aromatic rice line Jia 58: 199 plants showed the expected results, while the remaining 1 exhibited two fluorescent signal peaks-suggesting that it may be a heterozygous individual. Using the KASP marker, we performed genotyping analysis on F₇ progeny individuals derived from the cross between Yexiang (aromatic line) and Yuenongsimiao (non-aromatic line). Combined with phenotypic observations, we successfully screened out an elite aromatic line named Zhexiangzhenhe, which not only possesses aroma but also maintains superior agronomic traits. <b>Conclusions</b>: The multiplex-ready PCR assay and KASP markers facilitate high-throughput genotyping in large-scale breeding populations, providing breeders with a rapid and efficient selection tool to accelerate aromatic trait improvement in rice.