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PubMed2026

GWAS-based identification of a candidate gene and development of a predictive KASP marker for seed protein and oil contents in soybean.

BACKGROUND: Soybean [Glycine max (L.) Merrill] is one of the most widely cultivated crops worldwide. Its seeds contain about 40% protein and 20% oil, serving as essential nutrient sources for humans. Given the nutritional importance of seed protein and oil, identifying genes that regulate their levels is crucial for improving soybean seed quality. OBJECTIVE: This study aimed to identify genetic factors associated with seed protein and oil content using a genome-wide association study (GWAS). METHODS: Seed protein and oil contents were quantified in 192 soybean mutant accessions in a mutant diversity pool (MDP), and GWAS was conducted using 17,631 SNPs filtered from genotyping-by-sequencing. Expression of a candidate gene was examined across seed developmental stages (R5 to R7), and a significant SNP was converted into a Kompetitive Allele-Specific PCR (KASP) marker for validation. RESULTS: GWAS detected significant SNPs associated with seed protein and oil content. Chr20_7635098 was identified as a nonsynonymous SNP located in the exon of Glyma.20g042400. This gene showed differential expression across seed developmental stages between mutant accessions with contrasting protein and oil contents. The KASP marker for Chr20_7635098 was validated using the MDP and six domestic soybean cultivars showing predictive accuracies of ≥ 80.50% for protein content and ≥ 61.18% for oil content. CONCLUSION: Overall, this study identified a candidate gene linked to both seed protein and oil content, providing valuable insights for molecular breeding strategies aimed at efficiently improving these nutritional traits.

Glycine max↗

PubMed2026

Development and identification of KASP-SNP markers correlated with Aeromonas hydrophila resistance traits in blunt snout bream (Megalobrama amblycephala).

The blunt snout bream (Megalobrama amblycephala) is an economically important freshwater fish species. However, it is highly susceptible to Aeromonas hydrophila infection, especially in intensive pond aquaculture in China. Molecular marker-assisted selection provides an efficient approach for breeding disease-resistant varieties; however, the key genes or molecular markers linked to A. hydrophila resistance remain scarce in this species. A 436 differential SNP sites with disease-resistant were screened on basis of whole-genome resequencing. Then, a high-throughput genomic KASP genotyping technique was utilized to discover favorable genes and SNP sites associated with A. hydrophila resistance. A total of 46 KASP markers were successfully developed with an accuracy of 92 %. These markers were used to genotyping 120 blunt snout bream individuals. Through trait correlation analysis and general linear models (GLM), five SNPs significantly (P < 0.05) associated with resistance to A. hydrophila were identified and mapped to five candidate genes (btnl2, cfhr2, slc47a1, neu3, nlrp1). Survival rate of individuals carrying the dominant genotype demonstrated an average survival rate of 81.39 %, which represents a 69.35 % increase in comparison with that of 48 % in total population. This effect was validated in an external population of 100 fish. These findings identify key genetic markers associated with A. hydrophila resistance and provide a direction for elucidating the underlying molecular immune mechanisms, thus establishing a genetic foundation for future breeding strategies.

Cyprinidae↗

PubMed2026

Genome-wide association study and KASP development for growth and leaf traits in Populus deltoides.

BACKGROUND: Populus deltoides is a valuable timber species of considerable importance in the study of forest genetic breeding. However, its genetic improvement continues to rely predominantly on conventional selection and hybridization strategies hampered by long breeding cycles and limited efficiency. RESULTS: A total of 209 P. deltoides accessions were genotyped using a 60K SNP (Single nucleotide polymorphism) liquid array. Following quality control, 46,031 high-quality SNPs were screened and analyzed alongside 15 phenotypic traits in a genome-wide association study (GWAS), which identified 219 SNPs significantly associated with the traits. After further screening and annotation, a final set of 57 target SNPs and 77 candidate genes was obtained. Using kompetitive allele-specific PCR (KASP) assays, we successfully developed 48 polymorphic KASP markers. Of these, 25 markers exhibited significant phenotypic differences (p < 0.05) across genotype groups. CONCLUSIONS: These 25 KASP markers can serve as reliable and practical tools for phenotype-assisted selection, providing efficient molecular resources for accelerating genetic improvement and marker-assisted breeding in poplar.

Populus↗

PubMed2026

A transcriptome-wide approach for rapid pathotype discrimination of Puccinia striiformis f. sp. tritici in north-western India.

Stripe rust of wheat caused by Puccinia striiformis f. sp. tritici (Pst) remains a major constraint to wheat production in India due to the rapid evolution and frequent emergence of virulent pathotypes. Rapid and reliable discrimination of Pst pathotypes is essential for effective resistance deployment and surveillance. In the present study, transcriptome-wide simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs) were exploited to develop and validate molecular markers for pathotype-specific detection of Pst pathotypes prevalent in North India (110S119, 238S119, 46S119, 110S84 and 78S84). Microsatellite mining from 6103 core orthologous clusters comprising 51,127 transcripts mined 14,634 SSR loci, from which 93 primer pairs were synthesized. However, only three SSR markers exhibited polymorphism indicating limited discrimination potential of expressed sequence-derived (EST) SSRs for pathotype differentiation. In contrast, SNP discovery through stringent variant calling and filtration yielded 186 pathotype-specific homokaryotic SNPs, of which 56 high-confidence loci were selected for Kompetitive Allele-Specific PCR (KASP) assay development. A total of 48 KASP markers were synthesized and 14 demonstrated clear pathotype- or cluster-specific polymorphism representing substantially higher resolution than SSR markers. The high SNP-to-KASP conversion efficiency (~ 95%) and reproducible fluorescence-based clustering emphasize the robustness of KASP assay. Comparative evaluation revealed that SNP-based KASP markers provide superior discriminatory capacity for closely related Pst pathotypes and represent a promising complementary molecular approach for rapid identification of predominant Indian Pst pathotypes. The validated marker panel developed in this study can complement conventional virulence phenotyping and field pathogenomics approaches for surveillance of currently known pathotypes, while continued refinement may accommodate future changes in pathogen populations.

India↗

PubMed2026

Cloning of two Hsp70 genes and association analysis between SNP haplotypes and high temperature tolerance trait in red swamp crayfish (Procambarus clarkii).

Aquaculture is suffering the challenge from high temperature climate. Two Hsp70 genes, PcHsp70-1 and PcHsp70-2, as key genes involved in the high temperature tolerance of red swamp crayfish (Procambarus clarkii) were identified and cloned in this study. Their molecular features and expression patterns were characterized, revealing the distinct tissue-specific upregulation expression under high temperature stress (33 °C). Two SNPs, PcHsp70-1 (SNP258) and PcHsp70-2 (SNP555) were examined to associate with high temperature tolerance in three populations (n = 675). The genotypes of PcHsp70-1-SNP258 (GA) and PcHsp70-2-SNP555 (TT) were significantly associated with stronger high temperature tolerance. Notably, individuals carrying the haplotype of Hap I (GG + TT) showed a survival rate exceeding 70% under high temperature stress, whereas, the Hap VIII (AA + CT) showed it at 5.2%. RNA interference of PcHsp70-1 resulted in a significant decrease expression of the gene GSH-Px and its encoding protein (glutathione peroxidase) activity, and damage in intestinal tissue under high temperature stress. The transcriptome result revealed that PcHsp70-1 participates in regulation of the pathways related to cytoskeletal construction, immune response, apoptosis, and antioxidant defense. These findings indicate that PcHsp70 genes are crucial for the cellular stress response under high temperature stress. The developed Kompetitive Allele Specific PCR (KASP) markers provide valuable tools for the marker-assisted selection of high temperature tolerant crayfish varieties, supporting the sustainable development of aquaculture under the challenge of global warming.

Animals↗

PubMed2026

Identification and characterization of PsFwC9 conferring Fusarium wilt resistance in pea.

Pea (Pisum sativum L.) is one of the most important edible legumes in China, with both planting area and total yield ranking among the highest in the world. Fusarium wilt, caused by Fusarium oxysporum f. sp. pisi (Fop), is a severe factor limiting pea production. The deployment of resistant pea cultivars is the most effective and sustainable strategy for controlling this disease. In the present study, a novel resistance gene PsFwC9, conferring resistance to Fop race 5, was identified in the resistant pure line Chengwan 9-8 (CW9-8), and its candidate gene Psat4g213640 was characterized and functionally validated to be associated with disease resistance. Genetic analysis of the F₂ population derived from the cross between the resistant parent CW9-8 and the susceptible parent Chengwan 9-1 (CW9-1) revealed that PsFwC9 was controlled by a single dominant gene. Based on whole-genome resequencing, bulked segregant analysis sequencing (BSA-seq) and fine mapping, PsFwC9 was localized to an 817.06-kb region on chromosome 4 (i.e. linkage group IV, chr4LG4), flanked by KASP markers A016508 and A016511, and co-segregated with four markers. Haplotype analysis revealed that only the marker A016615 was significantly associated with Fusarium wilt resistance, and this marker was designated as a diagnostic marker for PsFwC9. Marker A016615 was located at 425 699 725 bp on chr4LG4, corresponding to the 277 bp within Psat4g213640, where a 'A/G' single-nucleotide polymorphism caused an amino acid substitution leading to an alteration in protein structure; therefore, Psat4g213640 was identified as the PsFwC9 candidate gene. Quantitative real-time PCR analysis showed no significant difference in the expression levels of Psat4g213640 between CW9-8 and CW9-1. Overexpression of the candidate gene Psat4g213640CW9-8 in the hairy root system significantly enhanced the resistance of CW9-1 to Fusarium wilt, whereas RNA interference-mediated silencing of Psat4g213640CW9-8 reduced the resistance of CW9-8, indicating that Psat4g213640CW9-8 played a crucial role in pea resistance to Fusarium wilt. In addition, subcellular localization showed that the protein encoded by Psat4g213640 was targeted to the endoplasmic reticulum. Collectively, these findings not only enriched the gene resources for disease resistance in pea and provided an important foundation for elucidating the molecular mechanism of PsFwC9-mediated resistance, but also provided important technical support for the practical application of molecular breeding for disease resistance in pea.

Journal Article↗

PubMed2026

Genome-wide identification of the carotenoid cleavage dioxygenase gene family in wheat and analysis of the TaDREB-7A-TaNCED9a regulatory module conferring drought tolerance.

Carotenoid cleavage dioxygenases (CCDs) play critical roles in plant growth, development, and abiotic stress responses, yet their genome-wide identification and drought response mechanisms remain unexplored in wheat. In this study, 34 TaCCD genes were identified in wheat, distributed across 15 chromosomes and phylogenetically classified into five subfamilies. Gene structure analysis indicated that members within each subfamily shared conserved motifs and similar intron-exon arrangements. Cis-regulatory element analysis suggested the potential roles of these genes in stress adaptation, developmental processes, and hormone signaling. Moreover, prediction of tertiary structures and protein-protein interactions revealed unique structural features and potential interacting partners of the TaCCD proteins. In addition, TaNCED9a, a member of the TaCCD family, showed the highest transcript level in wheat roots among all detected TaCCD genes and was significantly induced by drought stress. Subcellular localization assay indicated that TaNCED9a was located in chloroplasts. Downregulation of TaNCED9a expression led to reduced drought resistance in wheat, accompanied by an accumulation of reactive oxygen species and a decrease in endogenous abscisic acid levels. Using yeast one-hybrid, dual-luciferase, and tobacco transient co-expression assays, the upstream regulatory factor TaDREB-7A was identified, which can regulate the expression of TaNCED9a. Additionally, a KASP molecular marker was developed to identify the superior haplotype TaNCED9a-HapI, which exhibited a significantly higher germination rate compared to TaNCED9a-HapII under drought conditions, and was predominant in wheat. These results offer valuable insights into the TaCCD gene family's response mechanisms to drought stress in wheat, simultaneously identifying promising genetic resources for enhancing drought tolerance through molecular breeding.

CCD↗

PubMed2026

Development of molecular markers associated with saline-alkali tolerance in rapeseed (Brassica napus L.).

A total of 947 saline-alkaline tolerance-related molecular markers and a 5K cGPS genotyping chipwere developed, providing practical tools for marker-assisted selection and molecular design breeding of saline-alkaline-tolerant rapeseed. Rapeseed (Brassica napus L.) has relatively strong tolerance to saline-alkaline stress and shows great potential for the sustainable utilization and improvement of saline-alkaline soils. However, the breeding of highly tolerant cultivars still mainly depends on conventional hybridization combined with phenotype-based selection, which constrains breeding efficiency. In this study, previously reported saline-alkaline tolerance-related genes from rapeseed, rice, maize, wheat, sorghum, and Arabidopsis were collected. Candidate gene-based association analysis enabled the development of molecular markers and a genotyping chip. A total of 483 significantly associated genes were identified, among which 355 genes contained favorable haplotypes. Molecular markers were successfully developed for 275 genes, including 746 KASP and 201 InDel marker pairs, and four marker pairs were randomly selected for validation. In addition, a 5K cGPS liquid-phase chip (HZSW-cGPS-BRNAP-04), was developed and showed a high call rate and excellent reproducibility in genotyping. These markers and the chip are expected to improve the breeding efficiency of saline-alkaline-tolerant rapeseed cultivars. Overall, this study provides useful tools for early-generation evaluation and marker-assisted selection (MAS), and provides a foundation for molecular design breeding of saline-alkali-tolerant rapeseed.

Brassica napus↗