Use of near-isogenic lines to precisely map and validate a major QTL for grain weight on chromosome 4AL in bread wheat (Triticum aestivum L.).

This study precisely mapped and validated a major quantitative trait locus (QTL) on chromosome 4AL for thousand-grain weight in wheat using multiple near-isogenic lines. Thousand-grain weight (TGW) is an essential yield component. Following the previous identification of a major QTL for TGW within the interval of 15.7 cM (92.7-108.4 cM) on chromosome 4AL using the Nongda3338 (ND3338)/Jingdong6 (JD6) doubled haploid population, the aim of this study was to perform more precise mapping and validate the genetic effect of the QTL. Multiple near-isogenic lines (NILs) were developed using ND3338 as the recurrent parent through marker-assisted selection. Based on five independent BCF segregating populations derived from BCF plants with different heterozygous segments for the target QTL site and the results of genotyping analysis performed using the Wheat660 K SNP array, it was possible to delimit the QTL region to a physical interval of approximately 6.5 Mb (677.11-683.61 Mb, IWGSC Ref Seq v1.0). Field trials across multiple environments showed that NILs had a consistent effect on increasing the TGW by 5.16-27.48% and decreasing the grain number per spike (GNS) by 3.98-32.91% compared to the corresponding NILs, which exhibited locus-specific TGW-GNS trade-offs. Moreover, by using RNA sequencing (RNA-Seq) of whole grains at 10 days after pollination stage of multiple NILs, we found that differentially expressed genes between the NIL pairs were significantly enriched for cell cycle and the replication of chromosome-related genes, hence affecting cell division and cell proliferation. Overall, our results provide a basis for map-based cloning of the major QTL and determining the mechanisms underlying TGW-GNS trade-offs in wheat, which would help to fine-tune these two components and maximize the grain yield for breeders.