Analysis of Grain Protein Accumulation Differences and Screening of Associated Candidate Genes in Spring Wheat

Grain protein content in wheat is a key trait determining its nutritional and processing quality. Identifying key gene modules and core genes regulating its dynamic accumulation provides a theoretical basis for quality breeding. Using strong-gluten wheat variety Xinchun 48 (XC48) and medium-gluten wheat variety Humai 14 (HM14) as materials, grains were collected at 10, 15, 20, 25, and 30 days after anthesis for measurements of crude protein and soluble protein contents, followed by transcriptome sequencing. Weighted gene co-expression network analysis was employed to screen gene modules significantly associated with the target traits. Subsequently, differential gene expression analysis, functional enrichment analysis, and protein-protein interaction network analysis were integrated to identify core genes from the key modules. Phenotypic analysis revealed that the crude protein content of XC48 consistently exceeded that of HM14 from 10 to 30 days after anthesis, with statistically significant differences observed between 10 and 25 days after anthesis. In contrast, the soluble protein content was higher in HM14 during the period of 10 to 25 days after anthesis. However, this trend reversed at 30 days after anthesis, at which point XC48 exhibited a significantly higher soluble protein content than HM14. Transcriptome analysis identified a total of 27,499 differentially expressed genes, with expression peaks observed during the early and late developmental stages. Notably, transcription factor families such as ERF, MYB, NAC, and bZIP were prominently represented among these genes. Through WGCNA analysis, a MEgreen module exhibiting a highly significant positive correlation with protein content was identified. Based on protein-protein interaction network analysis, 14 core genes were screened. Further integration of expression patterns and functional annotations led to the identification of five key candidate genes: TaPP6R-C,TaNRPB11,TaRPS15A,TaRBX1and TaRPS29-like. qRT-PCR validation demonstrated that the expression dynamics of the five screened genes were highly consistent with the grain protein accumulation pattern. The findings provide a crucial theoretical foundation for further elucidating the molecular regulatory network underlying wheat grain protein formation, subsequent gene function validation, and breeding applications.