Fine mapping and candidate gene analysis of qKnps-2A for kernel number per spike in wheat

Kernel number per spike (KNPS) is a crucial determinant of wheat yield. Previous studies identified qKnps-2A as a major stable QTL for KNPS on chromosome 2A using a recombinant inbred line (RIL) population derived from a cross between the semi-winter wheat cultivar Kenong 9204 and the winter wheat cultivar Jing 411. However, fine mapping and gene cloning of this locus have not been reported.

Using InDel markers developed from parental genomic data, the near-isogenic line (NIL) pairs and 11 key recombinants were identified from residual heterozygous line-derived segregating populations, delimiting qKnps-2A to a 5.13 Mb interval (616.30-621.43 Mb) harboring 50 high-confidence genes. Integrated transcriptome sequencing, expression profiling across tissues and developmental stages, sequence comparison, and haplotype analysis identified TraesCS2A03G0902100, encoding a zinc knuckle protein, as the most likely candidate gene. Near-isogenic line analysis revealed that the Kenong 9204 allele significantly increased spikelet number per spike (2.94%), floret number per spike (10.51%), and KNPS (14.37%), resulting in a 7.32% yield increase, alongside improvements in grain quality traits. Phytohormone analysis showed the Kenong 9204 allele was associated with elevated IAA and IAA/ABA ratios during early spike development and reduced ABA during late development, suggesting hormone-related physiological changes linked to improved grain set. Haplotype analysis of 314 global accessions showed that Hap-KN9204 was significantly associated with increased KNPS, grain compactness, grain setting, and grain width. This superior haplotype predominates in Asia, Europe, and North America, and is suggestive of positive selection during Chinese wheat breeding, increasing from 30.00% in landraces to 59.62% in modern cultivars.

This study fine-mapped qKnps-2A and identified TraesCS2A03G0902100 as the most likely candidate gene, which was associated with altered phytohormone profiles during spike development, suggesting physiological changes linked to enhanced yield. The superior Hap-KN9204 haplotype appears to have been favored by positive selection in Chinese wheat breeding, establishing qKnps-2A as a valuable target for yield improvement.