Search Results (204)

Ongoing need for chromosome microarray analysis (CMA) characterization prompted the description of all 16,138 copy number variants (CNVs) found in 3832 patients studied from 2009 to 2024, 92% of them with developmental disabilities and/or autism. Detailed reporting shows the overlap of variants qualified as benign (15,083 CNVs, sizes 0.1 Kb–3 Mb) or of uncertain significance (216 CNVs, sizes 11 Kb–20 Mb) with pathogenic CNVs (836, 11 Kb–31 Mb), which are emphasized in most studies. Further distinguishing pathogenic CNVs were 88 recurring microdeletion/duplications and 86 in single patients, with all of the former and 66 of the latter having previous syndrome associations. Diagnoses were provided in 749 (20% of) patients, increasing to 21% among the 2470 patients (2015–2024) with their karyotypes recorded. Diagnoses included 61 known chromosomal syndromes, with CMA confirming or clarifying the abnormal karyotype in 187 (7.6%) or 55 (2.2%). The 90 microdeletions averaged 6439 kb in length (with chromosomes 6, 8, 17, and 22 accounting for most cases), while the 90 microduplications averaged 6895 kb (with chromosomes 8, 14, 17, 22, and X accounting for most cases). Together, these represent an average imbalance of 798,000 nucleotides per patient (0.75% of their genome). Continued reporting that match detailed CNV findings with patient profiles, especially symptom spectra, is needed to optimize CMA potential for presymptomatic diagnosis and therapy. Full article

Seed coat color is an important quality trait in mung bean (Vigna radiata) and is closely associated with seed appearance, commercial value, and phytochemical composition. To investigate the genetic basis of black seed coat formation, six F₂ populations were derived from reciprocal crosses between the black-seeded accession LZL115 and the green-seeded accession LZL156. Among 755 F₂ plants, 559 produced black-coated seeds and 196 produced green-coated seeds, conforming to a 3:1 segregation ratio (χ² = 0.37, p = 0.54). These results indicated that black seed coat color was dominant and consistent with the control by a single dominant locus, designated VrScL115, in the LZL115 × LZL156 genetic background. Bulked segregant analysis sequencing (BSA-seq) initially mapped VrScL115 to an approximately 2.90 Mb region on chromosome 4. Using newly developed KASP markers and recombinant screening in expanded F₂ populations, the locus was further delimited to a 121.79 kb interval between markers LS_K3333 and LS_K3379. Of the 11 annotated genes within this interval, LOC106758748 was the only gene containing high-confidence coding-sequence variants between the parents. This gene encodes a putative R2R3-MYB transcription factor homologous to MYB90. Comparative sequence analysis identified several allelic variants potentially associated with black seed coat color, and protein structure prediction indicated local structural differences between the parental proteins. LOC106758748 showed consistently higher expression in the developing seed coats of LZL115 than in those of LZL156 at 10, 15, and 20 days after pollination, with expression peaking at 15 days. Haplotype analysis of 246 mung bean accessions showed that the LZL115-associated allele combination at LS_K3352, LS_K3365, LS_K3367, and LS_K3370 was present in 21 of 27 black-seeded accessions (77.8%) and absent from all 219 non-black accessions, corresponding to a specificity of 100% and a false-negative rate of 22.2%. These findings support LOC106758748 as the leading candidate gene for VrScL115; however, direct in vivo functional validation is still required to confirm its causal role in black seed coat formation. The four-marker combination may be useful for identifying germplasm carrying the LZL115-associated allele, although further validation in independent germplasm populations is required. Full article

During high-concentration ethanol fermentation, Saccharomyces cerevisiae often faces multiple stresses, such as high osmotic pressure, ethanol toxicity, and nutrient limitation. These factors collectively limited the production of ethanol. To identify novel targets related to fermentation performance, we employed SHPERM- bCGHR strategy (a marker free allele replacement strategy based on comparative genomics and homologous recombination). We replaced the endogenous PHO4 of the high-producing strain MF01 with the PHO4 allele from MC15, thereby constructing a novel engineered strain MF01-PHO4. Under low-phosphate conditions, compared with the wildtype strain, the PHO5/11/12 genes and ribosomal protein genes showed significant upregulation in MF01-PHO4. These changes were associated with enhanced phosphorus uptake and protein synthesis. Under high phosphate conditions, the PHO4 expression and glycolytic enzyme gene expression in MF01-PHO4 were both lower than MF01, indicating that the substitution of the PHO4 allele may be associated with the coordinated changes in phosphate signal-mediated carbon phosphorus metabolism. This study identifies PHO4 as a promising candidate target for improving high concentration ethanol fermentation efficiency. These findings provide a framework to understand the phosphate-dependent regulatory effects of PHO4 allelic variation and offer a transferable strategy for strain improvement. Full article

Plant genomes exhibit extensive structural diversity generated by large-scale genomic alterations, collectively known as structural variations (SVs). Unlike single nucleotide polymorphisms (SNPs) and small insertions/deletions (indels), SVs can reshape genome architecture through changes in sequence content, gene dosage, regulatory landscapes, and chromosome organization. Recent advances in long-read sequencing (LRS), pan-genome construction, and multi-omics technologies have greatly expanded our ability to identify and interpret SVs across plant species. In this review, we summarize recent progress in understanding the formation mechanisms, classification, and functional consequences of plant SVs. We discuss major sources of SV generation, including transposable element activity, non-allelic homologous recombination (NAHR), horizontal gene transfer (HGT), and genome restructuring following polyploidization. We further highlight how LRS and graph-based pan-genomes overcome limitations of traditional linear reference genomes and enable more comprehensive characterization of genetic diversity. Beyond variant discovery, we emphasize the importance of integrating genomic, transcriptomic, epigenomic, proteomic, metabolomic, and spatial omics datasets to decipher how SVs influence gene regulation and complex agronomic traits. We also discuss current challenges, including repetitive genomes, polyploidy, computational complexity, and translation of SV knowledge into practical breeding applications. Together, these advances establish SV-centered genomics as a critical framework for understanding plant genome evolution and accelerating precision crop improvement. Full article

Background/Objectives: Colorectal cancer (CRC) is molecularly heterogeneous, and the KRAS (Kirsten rat sarcoma viral oncogene homolog) genotype both governs eligibility for anti-EGFR (epidermal growth factor receptor) therapy and carries prognostic weight. Central Asian data are scarce: no Kazakhstani series has described the KRAS variant spectrum, linked it to survival, or reported mutation detection rates across specimen types and PCR platforms. Methods: We studied 332 patients with morphologically confirmed stage II–IV colorectal adenocarcinoma. KRAS status was determined in formalin-fixed paraffin-embedded (FFPE) tumor tissue by allele-specific real-time PCR (RT-PCR) covering six selected codon 12 and 13 variants. Overall survival (OS) was estimated by the Kaplan–Meier method for the whole cohort and with stage stratification. Separately, mutation detection rates were recorded in three non-overlapping groups of different patients: plasma RT-PCR on the Idylla platform (n = 30), plasma nanoplate digital PCR (dPCR) on QIAcuity One (n = 120), and a routine tissue RT-PCR series (546 evaluable of 550). Because these groups differed in patients, specimen type, and mutation panel, this comparison describes observed detection rates only and supports no inference about analytical sensitivity, specificity, or concordance. Results: KRAS was mutated in 149/332 tumors (44.9%); codon 12 supplied 80.5% of variants, led by G12D (32.2%), G12V (24.8%) and G13D (19.5%). Median OS did not differ between mutant and wild-type tumors (39.0 vs. 36.6 months; p = 0.40). Variant-level differences were directionally consistent, but none was significant, and all were exploratory and unadjusted for multiplicity. Observed detection rates were 40.3% for tissue RT-PCR (220/546), 13.3% for plasma RT-PCR (4/30; continuity-corrected p = 0.006 vs. tissue), and 50.8% for plasma dPCR (61/120; continuity-corrected p = 0.044, Pearson p = 0.034 vs. tissue). BRAF V600E was detected by plasma dPCR in 11/120 cases (9.1%). Conclusions: This first Kazakhstani series places KRAS frequency within the internationally reported range and shows that variant-level reporting reveals prognostic structure that a binary call conceals. The higher detection rate seen with plasma dPCR is hypothesis-generating, not evidence of platform superiority, and motivates a prospective paired-sample study with harmonized mutation panels. Full article

The elite maize inbred line Zheng58, female parent of the widely cultivated hybrid Zhengdan958, is renowned for conferring short stature and high-density tolerance. Despite its critical role in modern breeding, the genetic basis of its dominant dwarfing effect has remained elusive. In this study, we dissected the genetic architecture of six plant architecture traits in a recombinant inbred line (RIL) population derived from elite inbred lines Zheng58 and PH6WC. Phenotypic evaluations across four environments revealed high heritability with additive effects accounting for 45.5–64.6% of the total genetic variance. A total of 125 QTLs were identified for the traits in single-environment QTL mapping, and multi-environment analysis further detected 77 QTLs for the six traits. Notably, a major dominant-negative pleiotropic QTL was identified on chromosome 2 that consistently explains plant height (PH), plant height above ear (PHAE), and average internode length above ear (AILAE) across multiple environments. The Zheng58 allele at this locus acts dominantly to reduce plant height by approximately 8.7 cm, providing a genetic explanation for Zheng58’s characteristic dwarfing effect. Regional association mapping refined this QTL to a 351.9 kb interval harboring 13 candidate genes. Transcriptome analysis uncovered 205 differentially expressed genes (DEGs) within the QTL region, with only one DEG (Zm00001d005848) located in the pleiotropic hotspot QTL on chromosome 2. This candidate gene encodes a rhomboid protease homolog, and population-wide expression data showed significantly negative correlation with PH. Our study unveils the genetic mystery of Zheng58′s dominant dwarfing phenotype by pinpointing a pleiotropic QTL hotspot, offering a strategic target for molecular breeding of compact, high-density-tolerant hybrids in maize. Full article

Soybean seeds have long been regarded as “storehouses of high-quality proteins”. The breakdown of dietary proteins by digestive proteases is essential for achieving adequate protein digestibility in animals and humans. However, plants have evolved a diverse array of protease inhibitors that regulate or restrict protease activity. In soybean, these inhibitors are concentrated primarily within the 2S protein fraction. Trypsin inhibitors (TIs) of Kunitz trypsin inhibitor (KTI) and Bowman–Birk inhibitor (BBI) are the most impactful due to their strong anti-tryptic activity, which interferes with digestive proteases in humans and animals. Elevated TI levels render raw soybeans unsuitable for direct food or feed use unless thermal or processing inactivation treatments are applied. Elimination or reduction in KTI and BBI using classical and biotechnology-based breeding efforts is a promising strategy. Soybean germplasm harboring BBI null alleles has not been reported. Breeding only for low or null KTI content in soybean would not be sufficient for practical applications. Hybridizing IT105782 × PI 547656 and using the reported Kompetitive Allele-Specific PCR (KASP) markers represents an effective classical breeding strategy. Simultaneous CRISPR/Cas9-mediated knockout of key KTI and BBI genes is expected to enable the development of soybean lines with substantially reduced TI levels, an outcome that cannot be readily achieved through classical introgression of null alleles, as naturally occurring null BBI alleles have not yet been identified. Moreover, this approach avoids the linkage drag associated with donor-derived null KTI alleles. However, this approach remains challenging due to functional redundancy and compensatory effects among KTI and BBI family members, extensive sequence homology among KTI and BBI genes that complicates the minimization of off-target effects, and the genotype dependency of Agrobacterium-mediated soybean transformation. Microtiter plate AACCI/AOCS could be one practical option for measuring TIA in breeding programs in terms of precision. Potential trade-offs associated with reduced trypsin inhibitor levels, including possible effects on plant defense and stress resistance, should be investigated in future studies, as these aspects have received little attention in previous research. Full article

Background/Objectives: Single-stranded oligonucleotides (ssODNs) are used as donor templates for therapeutic gene editing by CRISPR-Cas9 cleavage and homology-directed repair (HDR). Although ssODN sequence fidelity is critical to the safety and efficacy of editing, standard quality control methods cannot resolve individual nucleotide errors. Methods: We performed deep sequencing of ssODNs from three manufacturers and amplicons from edited hematopoietic stem/progenitor cells. Results: We find that synthesis errors are present in all ssODNs tested at rates that vary more than two-fold among manufacturers, at positions that are dependent on sequence context. These synthesis errors are propagated into the genome by HDR at frequencies proportional to their abundance in the ssODN. In our sickle cell mutation correction protocol, the most prevalent SNEs are predicted to produce benign β-globin variants, while the less frequent frameshift deletions are predicted to generate β-thalassemia-like alleles. Conclusions: Current quality control standards are insufficient to detect these errors, and deep sequencing of ssODNs should be incorporated into regulatory submissions for clinical gene editing programs. Full article

In this study, we combined computational predictions with experimental validation as a hybrid strategy to explore whether the E protein of tick-borne encephalitis virus (TBEV) possesses autoimmune potential. Using in silico homology searches, we identified two viral epitopes (evglekl and vtgtqgt) within the TBEV E protein that share sequence identity with fragments of the human proteins DNAH7 and CSMD2. Antibodies against these epitopes were detected in the plasma of a subset of patients after natural TBEV infection. Notably, no such antibodies were found in recipients of the Tick-E-Vac vaccine, indicating that the current vaccine does not induce cross-reactive humoral responses to these epitopes. Further computational analysis predicted that these epitopes could be presented by HLA class II molecules (alleles DRB1*09:01 and DRB1*07:01), which are known to be associated with autoimmune pathologies. Molecular dynamics simulations confirmed stable binding of the peptides within the HLA grooves, with favorable binding energies. These findings suggest a possible involvement of T-helper cells in the autoreactive process. Natural TBEV infection can give rise to antibodies against epitopes homologous to human proteins, particularly in genetically predisposed hosts. While such homology alone does not predict the onset of autoimmune disease, it represents a risk factor. Full article

Tomato brown rugose fruit virus (ToBRFV) poses a major threat to global tomato (Solanum lycopersicum) production, as it can overcome conventional resistance genes that are effective against tobamoviruses. In this study, a multiplex CRISPR/Cas9 system was developed to target the SlTOM1 susceptibility gene family (SlTOM1a–d), which encodes host factors essential for tobamovirus replication. Six guide RNAs (gRNAs), designed following 12 off-target analyses, were assembled into a multiplex CRISPR/Cas9 construct using a Golden Gate cloning strategy and introduced into tomato genotypes through an Agrobacterium-based tissue culture transformation procedure. Although primary T₀ transformants exhibited chimeric mutation patterns, stable inheritance and segregation of edited alleles were confirmed in the T₁ generation. Sequence analyses identified diverse indel mutations across target loci, with SlTOM1d exhibiting the highest editing efficiency. Multiplex genome editing successfully generated single-, double-, and triple-mutant combinations, with higher-order mutants displaying the strongest tolerance phenotypes. Following mechanical ToBRFV inoculation, edited T₁ plants exhibited markedly reduced symptom severity, low viral accumulation, and improved fruit health compared to wild-type controls. RT-qPCR analysis further confirmed significantly reduced viral RNA levels, supporting a host-factor-mediated tolerance mechanism. Importantly, edited lines maintained normal growth and agronomic performance. Collectively, these findings demonstrate that multiplex CRISPR/Cas9-mediated targeting of SlTOM1 homologs represents a promising and practical strategy for improving ToBRFV tolerance in tomato breeding programs. Full article

Genome editing is widely used and conceptually simple, yet in practice, it is hindered by laborious workflows and high costs. These challenges stem from the difficulty of identifying and isolating cells that contain the desired user-defined modifications, a problem compounded by the wide variability in editing efficiencies across cell types. While homology-directed repair (HDR) provides a mechanism for precise genome modification following nuclease-induced double-strand breaks (DSBs), it is frequently outcompeted by the dominant mutagenic non-homologous end-joining (NHEJ) pathway in mammalian cells. Therefore, we developed a novel enrichment method, Essential HDRescue, to increase the frequency of HDR events at a target site by co-targeting an essential genomic locus. Using both intrinsic positive and negative selection at a common essential gene, we enabled enrichment of precise editing events at a second, unlinked target site. We demonstrated that co-targeting essential genes in cancer cell lines and iPSCs increased HDR rates without the need for an exogenous reporter or selective drug. Analysis of resulting clones revealed that Essential HDRescue produced up to a 6-fold increase in single-allele edits and an ~4-fold increase in homozygous edits relative to single-targeted controls. By harnessing the intrinsic cellular dependencies that arise from DSB repair at essential loci, Essential HDRescue offers a widely applicable method to improve precise genome editing outcomes in mammalian cells, leaving only a minimal, protein-silent scar at the essential gene. Full article

Rice blast, caused by the fungus Magnaporthe oryzae, is one of the most devastating threatening to global rice production. The narrow genetic background of modern rice cultivars exacerbates the shortage of durable resistance resources. In contrast, the wild rice species Oryza officinalis harbors abundant stress-resistance alleles and represents a valuable gene pool for identifying novel broad blast-resistance genes. The cloned resistance gene Pid2 is encoded in a receptor-like protein kinase conferring race-specific resistance against the M. oryzae isolate ZB15. In this study, three Pid2 homologs were isolated from O. officinalis. The special allele Pid2of-MD33 was transformed into “Yunjing 37(YG37), a blast-susceptible japonica rice cultivar” via Agrobacterium-mediated transformation. Quantitative real-time PCR analysis showed that Pid2of-MD33 was consistently expressed in various tissues of O. officinalis, with the highest transcript abundance detected in leaf mesophyll cells and plasma membranes. Inoculation with the M. oryzae isolate ZB15 revealed that transgenic YG37 lines expressing Pid2of-MD33 displayed significantly reduced lesion size and pathogen proliferation, suggesting recovered race-specific resistance. These results enrich the resistance gene resources for rice blast research and provide a promising candidate gene for rice blast resistance breeding. Full article

Robust genetic tools are a prerequisite for causal, perturbation-based tests of redox physiology in acetogens. Here we establish practical genetic entry points for Sporomusa sphaeroides DSM 2875 under strictly anaerobic handling. We first attempted genome editing via double-crossover allelic exchange targeting pyrF using a non-replicative pUC19-based knockout construct and 5-fluoroorotic acid counterselection. Diagnostic PCR identified ΔpyrF candidates with the expected size shifts, demonstrating that homologous recombination is technically feasible in DSM 2875; however, the ΔpyrF genotype exhibited severe growth defects and could not be stably maintained over repeated passages, indicating a key limitation of a pyrF-based workflow under our current conditions. We then evaluated multiple E. coli–anaerobe shuttle plasmids for introduction and maintenance. Among the tested vectors, pJIR751 reproducibly yielded erythromycin-resistant transformants after prolonged incubation and supported serial passaging on selective media. Plasmid retention was confirmed by diagnostic PCR from liquid cultures in all tested isolates. Importantly, this maintainable plasmid platform enables genetically grounded perturbation-and-rescue experiments under electrode- or Fe⁰-assisted conditions, allowing mechanistic hypotheses in bioelectrochemical acetogenesis to be tested causally rather than inferred from phenotypes alone. Together, these results define current practical boundaries for S. sphaeroides genetics and establish pJIR751 as a practical foundation for downstream genetic manipulation in bioelectrochemical studies. Full article

Tomato high pigment-2 (hp-2^dg, hp-2, and hp-2^j) mutant lines are characterized by mutations in the DE-ETIOLATED1 (SlDET1; Solyc01g056340) gene. SlDET1 is responsible for encoding a nuclear protein that acts as a negative regulator involved in light signaling, repressing photomorphogenesis. These tomato mutant lines are known for increased levels of antioxidant pigments in fruits, such as flavonoids and carotenoids, compared to the wild-type fruits. In this study, CRISPR/Cas9, followed by the non-homologous end joining mechanism of repair (NHEJ), was used to target the SlDET1 gene and investigate whether the effects of these mutations could mimic the effects of hp-2 mutant lines, improving the nutritional features of tomato fruits. Our results indicated that mutations generated by CRISPR/Cas9 NHEJ in the hp-2 and hp-2^j regions (exon 11) resulted in significant changes in the SlDET1 coding and protein sequences. These mutations caused a low survival rate of edited sprouts and regenerated plants with a very compromised capacity of allelic heritability of these mutations for the following generations. However, regenerated plants containing these site-specific mutations in the SlDET1 gene showed higher levels of phytochemicals in ripe fruits. Furthermore, these edited plants also showed an upregulation of structural genes involved in the synthesis of these biocompounds. Although the SlDET1 gene could be considered an interesting target gene for the nutritional improvement of tomato fruits, our results showed that mutations within its exon 11 are quite critical and can induce severe perturbations in plant physiology, with a compromised possibility to develop new stable edited lines. Full article

Gibberellins (GAs) play a crucial regulatory role in the growth and development of cotton (Gossypium hirsutum L.). Through bioinformatics analyses, we identified a total of 39 GA2ox genes (encoding gibberellin 2-oxidases) in the cotton genome, designated GhGA2ox1 to GhGA2ox39. Based on phylogenetic analysis, these genes were classified into five groups. We further examined their gene structures, conserved motifs, and chromosomal distributions, revealing that members within the same group shared similar structural and motif organizations. Collinearity and cis-element analyses provided important insights into the evolutionary history and regulatory potential of the GA2ox gene family in cotton. Notably, using nucleotide diversity (π) and population differentiation (F_ST) analyses across the entire family, we screened and identified nine candidate genes that underwent strong artificial selection during cotton domestication and improvement. Further haplotype-phenotype association analysis identified GH_D09G0919 (GhGA2ox31) as a key regulator of Plant Height (PH). To validate their regulatory roles, we analyzed the genotype distribution in accessions with extreme phenotypes. The results revealed divergent selection histories for these two loci: the favorable allele of GH_D01G0720 (GhGA2ox23) was already fixed in the tested population, whereas GH_D09G0919 maintained significant natural variation. Specifically, the Hap2 allele of GH_D09G0919 was significantly enriched in the shortest accessions compared to the tallest ones. Importantly, quantitative real-time polymerase chain reaction (qRT-PCR) analysis confirmed that the Hap2 allele drives significantly higher gene expression in leaves, suggesting that enhanced GA catabolism underlies the compact phenotype. Additionally, transcriptomic profiling revealed the tissue-specific expression patterns of candidate genes, implying their functional roles in development. Furthermore, functional validation using the Arabidopsis mutant of the homologous gene (AtGA2ox8) confirmed its conserved role in regulating plant height, as the mutant exhibited a distinct short-stature phenotype. These results uncover valuable genetic resources for molecular breeding to shape compact cotton architecture. Collectively, this study aims to analyze the evolutionary patterns of the cotton GA2ox gene family and to identify key genes that regulate plant height under artificial selection, providing theoretical support for molecular breeding of compact plant types. Full article