Allele mining and haplotype-based association studies for nitrogen stress-responsive gene Os06g0291500 in indica rice (Oryz sativa L.) panel

Improving nitrogen use efficiency (NUE) in rice is critical for reducing nitrogen-induced environmental pollution and enhancing yield. Identifying natural allelic variants associated with NUE-related traits and utilizing them in breeding programs is an important approach. In this study, Os06g0291500, a novel, unannotated candidate gene associated with NUE, was used. Allelic variations were analyzed in 91 diverse rice genotypes through Illumina NovaSeq 6000 sequencing of the gene (3,078 bp) and its promoter (2,000 bp) regions. Mutation analysis revealed ample genetic variation of the gene across a panel of rice genotypes under study. The released varieties were found to accumulate more mutations, indicating limited selection for this gene in breeding programs. The phenotypic study showed differential N-response traits among genotypes, but association analysis did not reveal significant genotype-phenotype correlations due to the low phenotypic variance explained (PVE). Consequently, haplotype-based analysis was employed, identifying seven haplotypes within the panel. Among these, Hap_5 and Hap_7 were identified as the best and poorest haplotypes, respectively, based on phenotypic performance and SSI scores. The results highlight the potential of Os06g0291500 for NUE improvement in rice through functional validation and haplotype-based breeding. This study provides valuable insights into the genetic regulation of NUE and underscores the importance of exploring natural allelic diversity to develop nitrogen-efficient rice varieties.