Search Results (4,525)

Somatic cell preservation is an effective strategy for conserving the genetic potential of endangered species. To contribute to the conservation of the Milu deer (Elaphurus davidianus), this study aimed to establish and characterize an immortalized skin fibroblast cell line (ML-iSFC). The cell line is based on fibroblasts from the skin tissue of a male fawn of Milu deer. Optimal culture conditions were determined by supplementing the culture medium with different growth factors, and immortalization was achieved through simian virus 40 large T antigen (SV40T) transduction. Optimal culturing conditions for the cells were determined by adding a range of growth factors. The cellular morphology, growth characteristics, and marker expression of the cells were further evaluated. Cell cycle and proliferation were assessed by flow cytometry and CCK-8 assays, respectively. Chromosomes were determined by karyotype analysis. The highest cell growth rate was observed when the culture medium was supplemented with 3 ng/mL of FGF2. The fibroblast-specific marker vimentin (VIM) was expressed in both ML-SFC and ML-iSFC, while the epithelial marker keratin 18 (KRT18) was weakly expressed in ML-SFC cells. Cell proliferation and cell-cycle analysis revealed that ML-iSFC exhibited a higher growth rate and greater vitality compared to ML-SFC. Karyotype analysis showed that ML-iSFC maintained the same chromosome number and morphology as ML-SFC. In summary, this study reports the successful construction of an immortalized fibroblast cell line from Milu deer, which will serve as a valuable tool for Milu deer conservation. Full article

Triploid breeding is a promising approach for developing seedless varieties, but the long juvenile phase of perennial fruit trees necessitates efficient early selection. In loquat (Eriobotrya japonica), a fruit crop with high demand for seedlessness, the relative contributions of hybridity, ploidy level, and parent-of-origin effects (POEs) to triploid seedling vigor remain elusive. To dissect these factors, we established a comprehensive experimental system comprising reciprocal diploid (2x), triploid (3x), and tetraploid (4x) hybrids from two genetically distinct cultivars. The ploidy, hybridity and genetic architecture of hybrid and parental groups were verified using flow cytometry, chromosome counting, newly developed InDel markers and genome-wide SNP analysis. Phenotypic evaluation of eight vigor-related traits revealed that plant height and soluble starch content were the most robust indicators of triploid heterosis in loquat. Notably, paternal-excess triploids [3x(p)] consistently outperformed all other groups. Quantitative analysis revealed POE as the main positive driver of triploid heterosis (+10.37% for plant height), far exceeding the negative impacts of hybridity (−12.75%) and ploidy level (−20.87%). These findings demonstrate that POE predominantly drives seedling vigor heterosis in triploid loquat. We propose a practical breeding strategy that combines prioritizing paternal-excess crosses with novel InDel markers for rapid verification of superior seedless progeny. Full article

The cereal cyst nematode, Heterodera avena, is responsible for substantial economic losses in the global production of wheat, barley, and other cereal crops. Extracellular enzymes, particularly those from the glycoside hydrolase 18 (GH18) family, such as chitinases secreted by Trichoderma spp., play a crucial role in nematode control. However, the genome-wide analysis of Trichoderma longibrachiatum T6 (T6) GH18 family genes in controlling of H. avenae remains unexplored. Through phylogenetic analysis and bioinformatics tools, we identified and conducted a detailed analysis of 18 GH18 genes distributed across 13 chromosomes. The analysis encompassed gene structure, evolutionary development, protein characteristics, and gene expression profiles following T6 parasitism on H. avenae, as determined by RT-qPCR. Our results indicate that 18 GH18 members in T6 were clustered into three major groups (A, B, and C), which comprise seven subgroups. Each subgroup exhibits highly conserved catalytic domains, motifs, and gene structures, while the cis-acting elements demonstrate extensive responsiveness to hormones, stress-related signals, and light. These members are significantly enriched in the chitin catabolic process, extracellular region, and chitinase activity (GO functional enrichment), and they are involved in amino sugar and nucleotide sugar metabolism (KEGG pathway enrichment). Additionally, 13 members formed an interaction network, enhancing chitin degradation efficiency through synergistic effects. Interestingly, 18 members of the GH18 family genes were expressed after T6 parasitism on H. avenae cysts. Notably, GH18-3 (Group B) and GH18-16 (Group A) were significantly upregulated, with average increases of 3.21-fold and 3.10-fold, respectively, from 12 to 96 h after parasitism while compared to the control group. Meanwhile, we found that the GH18-3 and GH18-16 proteins exhibit the highest homology with key enzymes responsible for antifungal activity in T. harzianum, demonstrating dual biocontrol potential in both antifungal activity and nematode control. Overall, these results indicate that the GH18 family has undergone functional diversification during evolution, with each member assuming specific biological roles in T6 effect on nematodes. This study provides a theoretical foundation for identifying novel nematicidal genes from T6 and cultivating highly efficient biocontrol strains through transgenic engineering, which holds significant practical implications for advancing the biocontrol of plant-parasitic nematodes (PPNs). Full article

Karyotype analysis and the investigation of chromosomal variations in Populus are challenging due to its small and morphologically similar chromosomes. Despite its utility in chromosome identification and karyotype evolutionary research, satellite DNA (satDNA) remains underutilized in Populus. In the present study, 12 satDNAs were identified from P. trichocarpa, and the copy numbers and chromosomal distributions of each satDNA were analyzed bioinformatically in the reference genomes of P. trichocarpa, P. simonii, and P. nigra. Ten satDNA probes for fluorescence in situ hybridization (FISH) were successfully developed and validated on chromosomes of P. xiaohei (poplar hybrid P. simonii × P. nigra). By integrating bioinformatic genomic satDNA distribution patterns with experimental FISH signals, we constructed a molecular karyotype of P. xiaohei. Comparative analysis revealed errors in current poplar genome assemblies. Comparative karyotype analysis of P. xiaohei and its doubled haploid (DH) lines revealed chromosomal variations in the DH lines relative to the donor tree. The results demonstrate that the newly developed satDNA probes constitute robust cytogenetic tools for detecting structural variations in Populus, while molecular karyotyping provides new insights into the genetic mechanisms underlying chromosome variations in P. xiaohei and the DH plants derived. Full article

Bacterial wilt, caused by Ralstonia solanacearum, is a major constraint to tomato production globally. To uncover resistance loci and develop efficient molecular tools for breeding, we conducted disease phenotyping over two growing seasons, which revealed consistent variation in resistance and moderate broad-sense heritability (H² = 0.22–0.28), suggesting a genetic basis. A genome-wide association study (GWAS) was performed on a diverse panel of 267 tomato accessions, evaluated against two R. solanacearum strains. A major resistance locus was identified on chromosome 12, with the strongest association observed at SNP S12_2992992, located within a gene encoding a leucine-rich repeat (LRR) receptor-like protein. Haplotype analysis indicated that the resistance-associated allele is relatively rare (~13.5%) in the population, underscoring its potential value in breeding programs. Functional validation in an F₂ population derived from a cross between the susceptible ‘Seedathip6’ and the resistant ‘Hawaii 7996’ confirmed that the TT genotype at S12_2992992 was significantly associated with enhanced resistance. A Kompetitive Allele Specific PCR (KASP) marker was developed for this SNP, facilitating cost-effective and high-throughput selection. Collectively, these findings establish S12_2992992 as a robust and functionally informative marker, offering a valuable tool for accelerating bacterial wilt resistance breeding in tomato through marker-assisted selection. Full article

Terpene synthases (TPS) facilitate terpenoid production, influencing the flavor, color, and medicinal properties of Crocus sativus (saffron), a triploid geophyte of significant commercial importance. Despite its importance, the CsTPS gene family remains poorly characterized, limiting genetic enhancements in saffron’s agronomic features. This research performed a comprehensive genome-wide analysis of CsTPS genes using genomic, transcriptomic, and in silico approaches. BLASTP and PfamScan discovered thirty CsTPS genes, demonstrating conserved TPS domains, varied exon–intron architectures, and chromosomal clustering indicative of tandem duplications. Phylogenetic research categorized these genes into five subfamilies (TPS-a to TPS-e), with the prevalence of TPS-a suggesting a role in sesquiterpene biosynthesis. RNA-seq data (PRJNA976833, PRJNA400472) revealed tissue-specific expression, with CsTPS1 and CsTPS5 expressed in reproductive tissues and CsTPS2 in vegetative tissues. Stress-responsive genes (CsTPS1, CsTPS4) exhibited upregulation in response to cold and pathogen stress, with cis-regulatory elements (e.g., ARE, ABRE) indicating hormone control. The in-silico validation of CsTPS1, chosen for its elevated GMQE score (0.89), included primer design, ePCR, and vector optimization for expression in Arabidopsis thaliana. This study elucidates the contribution of the CsTPS family to saffron terpenoid diversity, providing a foundation for enhancing flavor, yield, and stress tolerance through genetic engineering. Full article

Stomata on the leaves of wheat serve as important gateways for gas exchange with the external environment. Their morphological characteristics, such as size and density, are closely related to physiological processes like photosynthesis and transpiration. However, due to the limitations of existing analysis methods, the efficiency of analyzing and mining stomatal phenotypes and their associated genes still requires improvement. To enhance the accuracy and efficiency of stomatal phenotype traits analysis and to uncover the related key genes, this study selected 210 wheat varieties. A novel semantic segmentation model based on transformer for wheat stomata, called Wheat Stoma Former (WSF), was proposed. This model enables fully automated and highly efficient stomatal mask extraction and accurately analyzes phenotypic traits such as the length, width, area, and number of stomata on both the adaxial (Ad) and abaxial (Ab) surfaces of wheat leaves based on the mask images. The model evaluation results indicate that coefficients of determination (R²) between the predicted values and the actual measurements for stomatal length, width, area, and number were 0.88, 0.86, 0.81, and 0.93, respectively, demonstrating the model’s high precision and effectiveness in stomatal phenotypic trait analysis. The phenotypic data were combined with sequencing data from the wheat 660 K SNP chip and subjected to a genome-wide association study (GWAS) to analyze the genetic basis of stomatal traits, including length, width, and number, on both adaxial and abaxial surfaces. A total of 36 SNP peak loci significantly associated with stomatal traits were identified. Through candidate gene identification and functional analysis, two genes—TraesCS2B02G178000 (on chromosome 2B, related to stomatal number on the abaxial surface) and TraesCS6A02G290600 (on chromosome 6A, related to stomatal length on the adaxial surface)—were found to be associated with stomatal traits involved in regulating stomatal movement and closure, respectively. In conclusion, our WSF model demonstrates valuable advances in accurate and efficient stomatal phenotyping for locating genes related to stomatal traits in wheat and provides breeders with accurate phenotypic data for the selection and breeding of water-efficient wheat varieties. Full article

Low light intensity is a major abiotic stress that severely affects rice yields, particularly in India and Southeast Asia, causing yield reductions of 35–40% during the wet season compared to the dry season. Tolerant rice genotypes exhibit adaptive changes at anatomical, physiological, biochemical, and molecular levels under low-light stress, enabling higher yields compared to susceptible varieties. Our study identified 20 novel QTLs associated with grain yields and nine related traits under low-light and control (normal)-light conditions, using a recombinant inbred line (RIL) population derived from the cross between the low-light-tolerant variety Swarnaprabha and the low-light-susceptible variety IR8. Across the Kharif seasons of 2019 and 2021, 33 stable QTLs were identified, with 11, 13, and 9 QTLs specific to low-light, normal-light, and both conditions, respectively. Of these, Swarnaprabha contributed 28 QTLs, while five were contributed by IR8. Notably, the study identified 11 and 9 novel QTLs under low-light and both conditions, respectively. Three hotspot regions on chromosomes 1, 4, and 8 were identified. These regions harbored 10 novel QTLs and revealed twenty candidate genes, out of which three key hub genes, OsAUX1, OsSBDCP1, and OsNPF5.16, were identified. These hub genes are involved in hormone signaling, starch metabolism, and nitrogen metabolism, respectively. A comprehensive expression analysis of these genes indicated that they are linked to low-light tolerance, offering deeper insights into the genetic and molecular mechanisms underlying low-light resilience. These findings provide valuable genomic resources and potential markers for breeding programs for improving rice productivity under low-light conditions. Full article

β-D-xylosidases (BXLs) are pivotal enzymes in xylan degradation, playing essential roles in plant development and stress responses. In this study, we identified 29 GmBXL genes in soybean through homolog alignment. Phylogenetic analysis classified these genes into three groups, with Group III being legume-specific. The GmBXLs are unevenly distributed across 15 chromosomes, with their expansion driven by both tandem and segmental duplications. Conserved motif and domain analyses revealed functional conservation, particularly in family 3 of glycoside hydrolase domains. Promoter regions of GmBXLs are enriched with hormone-responsive and stress-related cis-elements, indicating their involvement in diverse biological processes. Tissue-specific expression analysis revealed differential GmBXLs expression across leaves, roots, flowers, and seeds, with GmBXL13 and GmBXL26 exhibiting notably high transcript levels in pods and seeds. Under salt stress, 26 GmBXLs exhibited significant expression changes, with 20 genes up-regulated in both leaves and roots, highlighting their roles in salt tolerance. These findings enhance our understanding of the evolutionary and functional characteristics of GmBXLs, providing valuable insights for molecular breeding of salt-tolerant soybean varieties. Full article

Background/Objectives: Synovial sarcoma (SS) is an aggressive soft-tissue tumor characterized by the chromosomal translocation t(X;18) (p11.2;q11.2), most commonly involving the fusion of the SYT gene on chromosome 18 with the SSX1 or SSX2 genes on chromosome X. This study aims to explore the clinicopathological and molecular characteristics of synovial sarcoma in a cohort of Moroccan patients. Methods: We analyzed 48 cases of synovial sarcoma using formalin-fixed, paraffin-embedded (FFPE) tissue samples. Histological grading was performed according to the FNCLCC system. Immunohistochemical staining was employed to detect cytokeratin (CK) and epithelial membrane antigen (EMA). Molecular analysis included fluorescence in situ hybridization (FISH) to identify SS18 gene rearrangements and reverse transcription–polymerase chain reaction (RT-PCR) to detect SYT-SSX fusion transcripts. Results: Among the cohort, 56% of cases showed SS18 gene rearrangements via FISH, while RT-PCR confirmed the presence of SS18-SSX1 and SS18-SSX2 transcripts in 60% and 32% of cases, respectively. The remainder was classified as undifferentiated sarcoma. Notably, no significant associations were observed between SYT-SSX fusion type and clinicopathological features. Conclusions: These findings underscore the importance of integrating molecular techniques for precise diagnosis in synovial sarcoma. The results align with global patterns, emphasizing the necessity for molecular testing to enhance diagnostic accuracy and informing potential therapeutic advancements. Full article

Although heterosis plays a crucial role in enhancing crop yield and stress resistance, its underlying genetic mechanism remains not yet fully understood. Previous studies have shown that heterosis tends to increase with greater genetic distance in the absence of reproductive isolation barriers. However, whether variation in parental genome size alone can generate heterosis under near-isogenic backgrounds has not been thoroughly explored. Here, we used a rapeseed double haploid (DH) inducer line to generate progeny from the Pol CMS three-line hybrid Rongyou 18 (RY18). Although the progeny maintained the same ploidy level as the parents, their genome sizes showed notable variation (818.99–1024.88 Mb). To eliminate genetic distance effects, multiple DH progeny carrying restorer genes were crossed as paternal parents with the female parent 0068A of RY18, creating novel F₁ hybrids. Using RY18 as the control, we observed a marked reduction in the genetic distance between the newly induced restorer line and the female parent (0068A). Correlation analysis further revealed a significant negative correlation (r = −0.310 *) between the paternal genome size and heterosis for thousand-seed weight (TSW). Furthermore, the genomic expansion in hybrid offspring relative to the male parent showed that significant correlations were observed between paternal genome size and heterosis over the standard for both TSW (r = 0.300, p < 0.05) and plot yield (r = 0.326, p < 0.05). Resequencing of high-and low-yielding F₁ hybrids identified SNP sites, indicating that under an identical genetic background, heterosis for yield was more pronounced on chromosome A and chromosome C04. The doubled haploid (DH) induction line facilitates the generation of parental lines with distinct genome sizes, potentially providing a potential novel approach for studying heterosis research in Brassica napus. Full article

The LHCB gene family plays a crucial role in light harvesting and photoprotection in plants by encoding key components of the photosystem II antenna complex. The LHCB genes are also involved in salt stress. In this study, we systematically identified and characterized 16 LbaLHCB genes in the economically important medicinal plant Lycium barbarum. Comprehensive bioinformatics analyses revealed that these genes are unevenly distributed across seven chromosomes, with notable gene clustering on chromosome 11. Phylogenetic analysis classified them into seven distinct subfamilies, with the LbaLHCB1 subfamily showing significant expansion through gene duplication events. qRT-PCR and transcriptome analyses revealed tissue-specific expression patterns, with LbaLHCB1.6 exhibiting preferential expression in developing fruits, suggesting its potential involvement in fruit development and quality formation. Under salt stress conditions, the LbaLHCB genes displayed dynamic temporal responses: LbaLHCB1.5 was rapidly induced during early stress (1–3 h), LbaLHCB7 reached peak expression at mid-phase (6–12 h), while LbaLHCB1.2 showed significant downregulation during late stress response (24 h). Promoter analysis identified multiple stress-responsive cis-elements, providing molecular insights into their regulation under abiotic stress. These findings significantly advance our understanding of the LbaLHCB gene family’s structural characteristics and functional diversification in L. barbarum, particularly in relation to photosynthesis regulation and stress adaptation. The study provides valuable genetic resources for future molecular breeding aimed at improving stress tolerance and fruit quality in this important medicinal crop. Full article

Inducible CRISPR interference (CRISPRi) systems were established in Lactococcus lactis using both plasmid and chromosomal approaches. Expression of nuclease-deficient Cas9 (dCas9) from Streptococcus pyogenes was placed under the control of the nisin-inducible promoter P_nisA, while sgRNAs were transcribed from the constitutive P_usp45 promoter. To monitor expression, dCas9 was fused with superfolder GFP. Plasmid-based constructs successfully repressed a luciferase reporter gene and silenced the gene of the major autolysin, AcmA, leading to the expected morphological phenotype. However, plasmid systems showed leaky expression, producing mutant phenotypes even without induction. Chromosomal integration of dCas9 reduced its expression level by approximately 20-fold compared with plasmid-based expression, thereby preventing leaky activity and ensuring tight regulation. This chromosome-based (cbCRISPRi) platform enabled controlled repression of the essential gene ybeY, which resulted in severe growth defects. Restoration of wild-type phenotypes was achieved by introducing a synonymous codon substitution in the sgRNA target region. Transcriptome analysis of ybeY-silenced cells revealed downregulation of ribosomal protein genes and widespread effects on membrane-associated proteins, ATP synthase subunits, and various transporters. These inducible CRISPRi platforms provide robust and tunable tools for functional genomics in L. lactis, particularly for studying essential genes that cannot be deleted. Full article

Spinach is a dioecious vegetable and an excellent model for investigating plant sex differentiation. Exogenous gibberellin treatment induced sepal hypoplasia and sex reversal, converting 42% of stamens into pistils in male plants. Transcriptome analysis identified 112 male-biased genes enriched in stamen and pollen development, while hormone profiling revealed coordinated changes in GA, cytokinins, auxin, jasmonic acid, and abscisic acid. Functional assays demonstrated that silencing SpAMS or SpPGIP caused extensive carpelization, and in situ hybridization localized their expression to developing anthers. Dual-luciferase assays confirmed that SpAMS directly activates the B-class gene SpPI, and genomic mapping placed SpAMS in the pseudo-autosomal region of the Y chromosome. These results indicate that GA disrupts hormonal homeostasis and anther wall integrity, while the SpAMS–SpPI pathway regulates tapetal development to maintain male identity. Our findings identify SpAMS as a key male-promoting factor in spinach and provide a framework for elucidating sex determination mechanisms in dioecious plants. Full article

Cellular organisms store heritable information in two forms, genetic and epigenetic, the latter being largely dependent on cytosine methylation (5mC). Chargaff’s Second Parity Rule (CSPR) describes the nucleotide composition of cellular genomes in terms of intra-strand DNA symmetry. However, it remains unknown whether DNA methylation patterns display intra-strand DNA symmetry. Computational analysis was conducted of the DNA methylation patterns observed in human cell lines and in tissue samples from healthy donors. Analysis of 5mC marks in mutually reverse-complementary pairs of short oligomers, containing CpG dinucleotide in the middle, revealed deviations from CSPR and methylation asymmetry that can be observed for two non-overlapping mirror groups defined by CpG methylation values. Deviations from CSPR, together with combinatorial probabilities of pattern distributions and computer simulations, highlight the non-random nature of methylation processes and enabled us to identify specific cell types as outliers. Further analysis revealed a compensatory methylation asymmetry that reduces deviations from intra-strand symmetry and implies the existence of strand-specific methylation during cell differentiation. Among six pairs of reverse-complementary tetranucleotides, four pairs with specific sequence motifs display pronounced methylation asymmetry. This mirror asymmetry may be associated with chromosome folding and the formation of a complex three-dimensional landscape. Full article