Novel Machine-Learning Models Outperform Noninvasive Tests in Metabolic Dysfunction-Associated Steatohepatitis With Fibrosis.

Citric acid is the major organic acid affecting citrus fruit taste, which varies widely in the citrus family. An acidless kumquat, a small-fruited citrus species (Citrus crassifolia), has been developed and has become popular on the market. To investigate the molecular basis of the acidless phenotype in Huapi (HP) kumquat, it was crossed with early-flowering, high-acid Hong Kong (HK) kumquat to generate an F1 population, providing a system to dissect the genetic and molecular basis of citric acid accumulation. Organic acid content exhibited a wide and continuous distribution across three consecutive years, which is consistent with quantitative inheritance. Bulked segregant analysis sequencing mapped a major locus, and integration with transcriptomic data identified the GRAS transcription factor SCARECROW-like protein 9 (ChSCL9) as a candidate major-effect gene, showing high expression in HP fruit and low expression in HK fruit. Subcellular localization confirmed that ChSCL9 is a transcription factor. Functional analyses-including CRISPR-Cas9 gene editing, overexpression in kumquat, and RNA interference (RNAi) in citrus juice sacs-demonstrated that ChSCL9 negatively regulates citric acid accumulation. Biochemical experiments showed that ChSCL9 directly represses the expression of the R2R3-MYB gene ChPH4 and the vacuolar P-ATPase gene ChPH5, both of which are key genes for vacuolar acidification, thereby inhibiting citric acid accumulation. These results identify ChSCL9 as a key regulator of citric acid in kumquat, reveal an upstream regulator of PH4, and provide targets for citrus flavor improvement and rational design for citrus breeding.

Abstract The application of spur-type varieties is crucial for achieving dwarfing and dense planting cultivation. However, the underlying mechanisms regulating the spur-type phenotype remain unclear. In this study, we created an F1 population by crossing ‘Hanfu’ (HF, spur-type) with ‘Wangxianghong’ (WXH, standard-type) and observed that the spur rate in F1 hybrid population followed a normal distribution. Though bulked segregant analysis (BSA) and RNA-sequencing analysis (RNA-seq), we identified 85 differentially expressed genes across three QTL regions were screened. Interestingly, small auxin-up RNA 14 (MdSAUR14) exhibited distinct expression patterns in ‘HF,’ ‘WXH,’ and their extreme hybrid populations, suggesting a close association between its expression levels and the spur-type phenotype. MdSAUR14-overexpressed apple lines exhibited greater plant height and increased average internode length compared to wild type (WT) plants. A double base substitutions (DBS) variation in the MdSAUR14 promoter is closely linked to the spur phenotype. Furthermore, the presence of the DBS impacts the binding ability of BRASSINAZOLE-RESISTANT 1 (MdBZR1), which subsequently influences MdSAUR14 activity. Our study elucidates DBS-related genetic variation and mechanisms underlying shoot elongation in spur-type varieties.

Genetic improvement of clove (Syzygium aromaticum), an economically important spice and medicinal crop, has been constrained by limited molecular resources. The recent availability of a reference genome enabled a chromosome-scale genome-wide characterization and development of simple sequence repeats (SSRs) in clove. An in silico survey of the 367.77Mb genome assembly identified 189,127 SSRs at an overall frequency of 514.25 loci/Mb. Mononucleotides (50.30%) and dinucleotides (40.68%) were the most abundant repeat types, exhibiting a strong A/T-rich bias and a notable scarcity of CG/GC motifs. Although SSRs were uniformly distributed at the whole-chromosome level (R² = 0.99 relative to chromosome length), high-resolution analysis revealed significant intra-chromosomal heterogeneity, including distinct hotspots (> 600 loci/Mb) and coldspots (< 400 loci/Mb). A total of 11,107 hypervariable Class I loci were identified, predominantly comprising dinucleotide repeats (92.3%), highlighting them as major contributors to SSR variability. From these loci, 28,147 high-quality primer pairs targeting Class I and II SSRs were developed and curated into a publicly accessible, searchable Clove SSR Marker Database. Experimental validation of 55 primer pairs across 19 germplasm accessions demonstrated high amplification success (96.4%) and polymorphism rates (90.6%). The validated markers revealed moderate genetic diversity (mean He = 0.22; PIC = 0.18), consistent with a relatively narrow genetic base in the analyzed germplasm. Phylogenetic analysis resolved the 19 accessions into distinct clades and identified a highly divergent accession with red-coloured flower buds (Acc. 9833). The practical utility of the validated markers was further demonstrated through hybrid authentication and confirmation of co-dominant Mendelian segregation in an F₂ population. Collectively, this validated SSR marker resource and database provide a valuable genomic resource for clove genetic studies, germplasm management, and future genetic mapping efforts.

Omics-assisted genetic dissection and coordinated improvement of apple flesh color, flavor, and fruit size.

Angular leaf spot (ALS), caused by Xanthomonas fragariae, is a bacterial disease that limits strawberry (Fragaria × ananassa) productivity worldwide. Although major resistance loci have been identified in wild Fragaria species, their introgression into elite germplasm remains constrained by linkage drag and inconsistent inheritance. To dissect the genetic architecture of ALS resistance, we evaluated a diverse panel of n=241 greenhouse-tested and n=468 field-tested strawberry accessions representing elite cultivars, breeding materials, and heirloom varieties of cultivated strawberry (F. × ananassa). Plants were evaluated under controlled inoculations and natural field infection using a standardized phenotypic scale and genotyped with 50K single nucleotide polymorphism array. Broad-sense heritability was moderate (H2=0.45), whereas narrow-sense heritability was low (h2=0.10), reflecting the contribution of nonadditive genetic effects such as epistasis. Genome-wide association studies (GWAS) identified two loci on chromosomes 1D and 2D, explaining 4%-5% of phenotypic variance, respectively. A segregating F1 population validated the 2D locus, confirmed by co-localization with the GWAS signal explaining 11.3% of the phenotypic variance. Two-dimensional genome wide scan in this F1 population revealed a significant epistatic interaction between locus on chromosome 2D and an additional locus on chromosome 6B, collectively explaining 27% of the phenotypic variance. Together, these results demonstrate that nonadditive effects in these populations control ALS resistance. Understanding this complexity provides a foundation for developing elite cultivars with durable resistance, such as UCD Royal Royce. Our findings underscore the need for predictive breeding strategies that capture epistatic and environmental variance to accelerate genetic gain for ALS resistance in strawberry.

Fine-tuning heading date is critical for optimizing yield, grain quality, and local adaptation in rice (Oryza sativa L.). QTL-seq (where QTL is quantitative trait locus), which integrates bulked segregant analysis with whole-genome resequencing, provides a rapid and cost-effective approach for identifying QTLs underlying complex traits. Shinpyeong and Haedeul are elite Korean rice cultivars that carry identical alleles at four major heading date genes (Hd1, OsPRR37, Ghd7, and DTH8), yet differ by approximately 7 days in heading under natural long-day conditions. To elucidate the genetic basis of this difference, we applied allele-informed QTL-seq to an F2 population derived from a cross between Shinpyeong and Haedeul. A single prominent QTL peak with the highest ΔSNP-index (where SNP is single nucleotide polymorphism) was consistently detected across both field and greenhouse environments and co-localized with Hd18, identifying it as the most likely causal gene. The underlying polymorphism was a previously reported missense A/G variant. Genotyping of the Hd18 A/G SNP using a dCAPS (derived cleaved amplified polymorphic sequence) marker revealed that F2 plants homozygous for the A allele headed 4.1 days earlier in the field and 6.2 days earlier in the greenhouse than those homozygous for the G allele. Analysis of 290 Korean rice cultivars further demonstrated that the Hd18 A allele was associated with approximately 2 days earlier heading under a nonfunctional hd1 background. Evaluation of two recombinant inbred line populations showed that Hd18 had minimal effects on major agronomic traits, while exhibiting background-dependent effects on specific traits. These results establish Hd18 as a minor effect modifier of heading date with utility for fine-scale phenological adjustment in rice breeding.

Maize (Zea mays L.) is a globally important crop. Understanding plant height and the underlying dwarfing genes is of critical importance for improving lodging resistance and optimizing plant architecture. In this study, a dwarf mutant designated CM17-2 was obtained by exposing the maize inbred line B73 to carbon ion beam irradiation. The mutant exhibited markedly reduced plant height, internode length, and ear height. Segregation analysis in the F2 population revealed a 3:1 ratio, indicating monogenic recessive inheritance. Using bulked segregant analysis (BSA), the causal locus was mapped to four intervals on chromosomes 2, 4, and 5. Exogenous GA3 treatment rescued the dwarf phenotype and restored plant height to near-wild-type levels, indicating GA sensitivity. Transcriptomic analysis following GA3 treatment identified 3292 differentially expressed genes (DEGs) between CM17-2 and B73. Among these, 1664 GA3-responsive genes were significantly enriched in the plant hormone signal transduction pathway. Integration of BSA-seq and transcriptomic data pinpointed 18 candidate genes on chromosome 4. Through gene annotation, four potential candidate genes likely associated with dwarfism were identified, and the functions of these candidate genes need to be validated in future experiments. The identification of these potential candidate genes lays a theoretical foundation for the subsequent functional analysis, gene cloning, and mechanistic studies of maize dwarfism-related genes.

Edible podded peas viz., snow pea (Pisum sativum var. saccharatum) and sugar snap pea (Pisum sativum var. macrocarpon) are valued for their tender, parchment-free pods and sweet flavor. In present study, morphologically 26 diverse genotypes were characterized based on pod colour, flower colour, seed colour and pod type. Among these, four morphologically and phenotypically diverse genotypes viz., 'Royal Snow', 'Punjab Mithi Phali-3', 'Honey Snap' and 'Mithi Phali' were identified and utilized for developing reciprocal segregating populations between snow and sugar snap peas (Mithi Phali × Honey Snap, Honey Snap × Mithi phali, Royal Snow × Punjab Mithi Phali-3, Punjab Mithi Phali-3 × Royal Snow) to study the maternal/cytoplasmic inheritance. Phenotypically first generation hybrids across all reciprocal crosses indicated dominance of purple flower colour over white, green pod colour over yellow and purple pigmented pods, brown/light brown seeds over green colour seeds and snow-type over sugar snap pods without showing maternal/cytoplasmic inheritance. Segregating F2 populations followed expected Mendelian inheritance with chi-square (χ2) analysis confirming 3:1 best fit ratio, indicating single dominant gene inheritance for these traits in edible podded peas. These findings provide valuable insights for the genetic improvement of edible podded peas.

Waterlogging, like other abiotic stresses, severely inhibits the growth and development of watermelon. However, few genes conferring waterlogging escape response have been identified in this crop to date. In this study, we performed bulked segregant analysis on a F2 population exhibiting significant variation in waterlogging-induced shoot elongation. A 6.59 Mb candidate region on chromosome 7 was identified to be associated with waterlogging-induced shoot elongation. Integrated resequencing, fine-mapping and transcriptome analyses indicated that Cla97C07G134400 (ClSCPL50) candidate gene associated with waterlogging-induced shoot elongation. These findings provide novel insights into the genetic basis of the waterlogging escape response in watermelon and identify a potential target gene for improving waterlogging escape response in other dryland crops.

The development of aromatic rice varieties requires precise identification of aroma-associated alleles and validation of hybridisation during breeding programs. The present study evaluates the efficiency of SSR marker RM190 in detecting aroma-linked polymorphism and confirming hybridity in rice. Crosses between aromatic (Bio Tulsi) and non-aromatic (BPT-5204, TMRV-1509) genotypes were analysed along with their F₁ and BC₁F₁ populations. DNA was extracted using the CTAB method, followed by PCR amplification and agarose gel electrophoresis. The marker RM190 exhibited clear polymorphism between parental lines and successfully identified heterozygous F₁ plants through the presence of dual bands. Segregation patterns observed in BC₁F₁ populations followed expected Mendelian ratios, confirming the reliability of marker-assisted selection. The study highlights RM190 as a robust molecular tool for rapid screening of aroma traits and hybrid validation in rice breeding programs.

Brown midrib (bmr) mutants are frequently associated with unfavorable agronomic traits. In this study, we identified a novel brown midrib mutant, bmr34, which exhibited distinct brown coloration in roots, stems, and leaf midribs. Although most classic bmr mutants show undesirable agronomic performance, this mutant displays altered lignin accumulation and has important potential for forage quality and biomass utilization, providing a key genetic resource for lignin regulatory research in sorghum. Compared to the wild-type, bmr34 showed no significant differences in five major agronomic traits; however, lignin content was significantly reduced. Bulked segregant analysis (BSA) using an F2 population derived from a cross between bmr34 and the wild type Tx623 mapped the candidate region to chromosome 4. Further sequencing analysis identified a single nucleotide substitution (C → T; reverse strand G → A) at position 5,731,348 within the 5′ splice site of the third intron of Sobic.004G071000 in the mapping interval. KASP marker analysis demonstrated complete co-segregation between the mutation site and the bmr phenotype. Sequence analysis also revealed that this G → A substitution resulted in aberrant splicing and a 33-bp insertion in the third exon, which introduced a premature stop codon. Notably, the normally spliced transcript still accounted for approximately 36.2% of total transcripts in bmr34, indicating partial retention of wild-type transcript processing. These results demonstrate that bmr34 represents a novel weak allele of Bmr6, providing new insights into splice-site mutations and their contribution to lignin biosynthesis regulation in sorghum.

An indel in the FAD5 promoter confers cold tolerance by enhancing transcriptional activation of unsaturated fatty acid synthesis in grapevines.

Stem lodging significantly reduces soybean yield stability, particularly under dense planting and intercropping systems. Stem breaking strength is a key component of lodging resistance, but its genetic basis remains incompletely understood. In this study, an F2 population consisting of 167 individuals derived from a cross between nanxiadou25 (NXD25, high stem breaking strength) and Shiyuehuang (SYH, low stem breaking strength) was analyzed using bulked segregant analysis with whole-genome resequencing (BSA-Seq) to identify loci associated with stem breaking strength. The trait showed broad quantitative variation in the F2 population, ranging from 20.1 to 673.7 N. Two extreme bulks were constructed using 30 plants with the highest values and 30 plants with the lowest values. QTL-seq detected 21 candidate intervals at the 95% confidence level, among which, three major loci on Chr07, Chr13, and Chr16 exceeded the 99% threshold and were designated qBR7.2, qBR13.1, and qBR16.1. By integrating large-effect SNP/InDel variation, marker development, RNA-seq profiling, and qRT-PCR validation, nine candidate genes were retained for further study, and three marker-linked genes were highlighted as high-priority candidates. RNA-seq identified 9617 differentially expressed genes between the two parents. In addition, three co-dominant InDel markers, Chr07_01, Chr13_17, and Chr16_83, showed phenotype-consistent polymorphism in extreme F2 individuals. These findings provide valuable loci, candidate genes, and molecular markers for soybean lodging-resistance breeding.

Barley leaf stripe (BLS) is a common fungal disease caused by P yrenophora graminea in hulless barley (Qingke). Once leaf stripe occurs, it will seriously affect the yield and quality of Qingke. This study utilized the disease-resistant variety Kunlun14 (KL14) and the susceptible variety Z1141 as materials to analyze the changes in the disease index, soluble protein content, relative conductivity, proline (Pro) content, malondialdehyde (MDA) content, and chlorophyll content (CC) in leaves before and after infection with barley leaf stripe (BLS), which is caused by P. graminea. After infection, both the disease incidence and severity in the Z1141 group were significantly higher than those in the KL14 group. BSR-seq analysis was performed on the resistant and susceptible bulks derived from the parents and the F2 population (Z1141 × KL14), and three candidate regions were identified: one locus (648,449,784 base pairs (bp)) and one interval (659.7-668.0Mb) on chromosome 5H, and one interval (12.1-16.3Mb) on chromosome 6H. A total of 3,126 differentially expressed genes (DEGs) were identified by Isoform sequencing (Iso-seq) using leaves of KL14 and Z1141 before and after P. graminea infection. An integrated analysis of BSR-seq and Iso-seq data, supplemented by qRT-PCR validation and functional annotation, pinpointed HvWAK and HvRGA as candidate genes for BLS resistance. Expression analysis demonstrated that the expression levels of both genes peaked at the week seven following P. graminea infection, and the timing of transcriptional activation in Z1141 preceded that in KL14. This study analyzed the resistance mechanisms of Qingke against P. graminea through both physiological response analysis and identification of disease resistance genes. The results of this study not only contribute to a deeper understanding of plant-pathogen interaction mechanisms but also establish a valuable genetic resource base for molecular marker-assisted breeding of BLS-resistant Qingke and the functional characterization of the key candidate genes HvWAK and HvRGA.

Sweetpotato (Ipomoea batatas (L.) Lam.) is an important multifunctional crop with great value in food supply, industrial processing and bioenergy utilization. Crude protein content (CPC) is a core target trait for sweetpotato quality breeding. To dissect the genetic basis of CPC and identify key candidate genes, we used an F1 population of 212 individuals. CPC was measured by near-infrared reflectance spectroscopy (NIRS) in 2020 and 2021, and QTL mapping was performed using a high-density SNP genetic linkage map. Candidate genes were explored via a genome-wide association study (GWAS), multiple-database functional annotation, and quantitative real-time PCR (qPCR) validation. The results showed that: (1) CPC in the population exhibited a continuous normal distribution with high inter-year stability, and phenotypic variation was mainly controlled by genetic factors; (2) one stable minor-effect QTL for CPC, qCPC09-1, was mapped to Chr09: 7906895–8614924 bp, explaining 5.7% of phenotypic variation; (3) GWAS detected no significant SNP loci, suggesting that CPC is regulated by multiple minor-effect genes; (4) genes within the qCPC09-1 interval were significantly enriched in three protein synthesis-related KEGG pathways: ribosome, nitrogen metabolism and ubiquinone and other terpenoid–quinone biosynthesis; (5) qPCR verified that itf09g13420 and itf09g13230 were upregulated in the low-CPC parent Yushu 10 and negatively correlated with CPC, while itf09g13550 was upregulated in the high-CPC parent Xin 24 and positively correlated with CPC. These three genes exhibited expression patterns highly consistent with phenotypic differences. This study provides a theoretical basis and technical support for molecular marker-assisted breeding and elite germplasm innovation in sweetpotato.

Local chicken breeds have long been subjected to unique geographical isolation and extreme environmental pressures, which have led to significant environmental adaptation. This provides a valuable opportunity to explore the mechanisms of adaptive evolution in domestic chickens and to develop rare genetic resources. However, the molecular genetic basis of these local breeds remains inadequately studied. In this research, we systematically compared the population genetic structure and selection signals between the Hotan Black Chicken (HT) from Xinjiang and the F2 hybrid population of Yeonsan Ogye × White Leghorn chickens (OLF) based on medium-density SNP chip data. Principal Component Analysis (PCA), Neighbor-Joining (NJ) phylogenetic trees, and ancestry inference methods revealed significant genetic differentiation between the two populations. HT exhibited a relatively homogeneous genetic background, while the F2 population showed a more complex genomic structure. By combining a sliding window approach with population differentiation index (FST), nucleotide diversity (π), and the ratio log2(π_HT/π_OLF), we identified selection signal regions across the whole genome, with 99 candidate genes for the OLF population and 36 for the HT population. The candidate regions in the HT population were enriched in pathways related to environmental adaptation, including cell adhesion, neurodevelopment, and immune regulation, while the F2 population showed significant enrichment in PI3K/AKT signaling and protein phosphorylation pathways related to production performance. This study not only elucidates the adaptive genomic features of local chicken breeds in extreme environments but also provides a genomic basis for the conservation and molecular breeding of local chicken genetic resources.

Allelic variation in the thioredoxin TRX-M4 influences carbon partitioning to control soluble sugar content in apple fruit.

Plant height is a key component of sunflower (Helianthus annuus L.) plant architecture. It strongly influences lodging resistance, mechanized harvestability, and yield stability. However, the genetic basis of plant height in sunflowers remains underexplored. This study aimed to develop an F2 population consisting of 715 individuals from a cross between the dwarf inbred line 150A and the tall inbred line PT326. Bulked segregant analysis coupled with whole-genome resequencing was employed to identify loci associated with plant height. Using three complementary analytical methods, a major quantitative trait locus, qPH15, was identified on chromosome 15. This locus was subsequently fine-mapped, using Kompetitive Allele Specific PCR (KASP) markers and recombinant screening in F2 and F3 populations, narrowing it to a 64.66-kb region containing three annotated genes. Among these, HanXRQr2_Chr15g0707451, which encodes an NAC transcription factor designated HaNAC7, was identified as the most promising candidate gene. Haplotype analysis of HaNAC7 across 148 sunflower accessions revealed 4 polymorphic sites defining 6 haplotypes with substantial differences in plant height. The shortest haplotypes, Hap2 and Hap3, were associated with reduced plant height and were predominantly found in Asian germplasm. These findings suggest that HaNAC7 is a strong candidate gene underlying qPH15 and provide useful molecular markers and favorable allelic resources for improving sunflower plant architecture.

BackgroundWater soaking (WS) is an economically important surface disorder in open-field production of strawberry (Fragaria × ananassa). The disorder, which impairs fruit quality and quantity, develops after exposure of maturing fruit to rain or high concentrations of water vapor. The susceptibility to WS differs among genotypes, suggesting a genetic component, with earlier studies establishing a close positive relationship between susceptibility to WS and the water permeance of the fruit skin. However, until now, no published studies on the underlying genetics of WS exist. Here, we aimed to identify genetic regions associated with WS and fruit skin permeability using a genetic mapping approach.ResultsA biparental F1 population was established by crossing two genotypes (201409 × 210706) exhibiting contrasting susceptibility to WS. The trueness-to-type of the progenies was confirmed by genetic fingerprint analysis with molecular markers. WS and permeance for water uptake were phenotyped over two seasons under two different growing conditions using laboratory-based incubation assays. Furthermore, the population was genotyped with the Axiom™ Strawberry FanaSNP 50K Genotyping Array to facilitate genetic mapping and QTL analyses. WS and skin permeance showed a clear segregation and were closely related. Multiple QTL regions were identified for WS and permeance, with one accounting for 16.7% to 28.6% of the phenotypic variation. ConclusionThe detection of several QTLs and the moderate broad-sense heritability of 0.49 suggest a polygenic background of WS. These findings provide the first steps towards understanding the genetic background of this trait, which will allow for developing tolerant cultivars for open-field production.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12870-026-08778-2.