Identification and validation of qPC-3A, a stable QTL for flour protein content in hard wheat (Triticum aestivum L.).

key messageA novel and stable QTL was identified for wheat protein content, independent of kernel traits, with a validated KASP marker for efficient breeding without yield penalties. Protein content (PC) is a critical determinant of wheat end-use quality. While NAM-B1 is a major gene regulating PC in wheat, its breeding applications are limited due to its wild origin and undesirable agronomic traits. This study aimed to identify a novel quantitative trait locus (QTL) for PC that is independent of yield-related kernel traits using a recombinant inbred line (RIL) population derived from the white hard wheat cultivars Jokyoung and Joongmo2008. Weak or no significant correlations were observed between PC and kernel traits, such as kernel number per spike, thousand-kernel weight, kernel length, kernel width, and kernel length-to-width ratio. A major QTL, qPC-3A, was identified on chromosome 3A with a logarithm of odds score of 8.2, a phenotypic variance explained of 22.2%, and 1.2 percentage point increase in PC contributed by Joongmo2008 over three years of field trials. The qPC-3A locus was independent of Glu-A1 and Glu-B1 alleles, and no QTLs associated with other kernel traits were detected on chromosome 3A. The qPC-3A interval was mapped to 0.76 Mbp, and a Kompetitive Allele-Specific PCR (KASP) assay was developed and validated. Evaluation of the RIL population and Joongmo2008-derived breeding materials confirmed that the KASP assay effectively distinguished high- and low-PC lines. Furthermore, qPC-3A demonstrated reliability across diverse breeding lines and multiple years of testing. The identification of this novel and stable QTL for PC, along with its associated KASP marker, provides a valuable genetic resource for wheat breeding programs aimed at improving protein content without compromising the grain yield.