NAM-BSA: A Bulked Segregant Analysis Method Using a NAM Population

Common gene mapping approaches mainly include QTL-mapping, BSA-seq (QTL-seq) and GWAS. BSA-seq is designed to facilitate the mapping of quantitative trait loci (QTL) in a cost-effective and high-efficiency manner. In order to accommodate the diverse species-specific traits and population genetic architectures, researchers have developed a series of tailored BSA methodologies. In this study, using six wild rices (Oryza rufipogon) as donors and the elite cultivated rice Youzhan 8 (YZ8) as the recipient, we constructed a BC₄F₈ population through hybridization and backcrossing. These six wild rice introgression lines together constitute a nested association mapping (NAM) population. Upon genotyping 1819 lines of the NAM population for the Sd1 gene, we found that among lines harboring the 383 bp deletion, 98.5-99.3% exhibited a low plant height (LP) phenotype, whereas 0.7-1.5% showed a high plant height (HP) phenotype. Based on this phenotypic segregation, we selected a total of 20 HP lines and 20 LP lines from six BC₄F₈ populations to form the H-bulk and L-bulk, respectively. Using BSA-seq, we identified a total of 33 significantly associated candidate intervals. One candidate interval located on chromosome 1 contains D18, a previously reported gene that regulates plant height. Furthermore, we preliminarily identified two major candidate genomic regions on chromosome 8 (4.60-5.76 Mb and 7.29-7.33 Mb). Integrating RNA-seq data, CAFRI-Rice online functional prediction and RT-qPCR validation, two key candidate genes, LOC_Os08g09900 and LOC_Os08g09000, were selected for subsequent functional characterization. The results of this study indicate that the NAM-BSA technology has great potential to detect QTL associated with complex traits, which can provide a novel technical strategy for the genetic dissection of complex traits in rice.