Sheng wu gong cheng xue bao = Chinese journal of biotechnology最新文献

Liquid-liquid phase separation (LLPS) is an important mechanism that drives intracellular biomolecules to form membraneless organelles, which plays an important role in regulating plant development and stress responses. In recent years, as the research on phase separation advances, remarkable progress has been achieved in the development and application of related technologies, which provides important technical support for the research in this field. From the basic concepts and principles of phase separation, this paper systematically summarizes the widely used labeling imaging techniques, computational prediction techniques based on machine learning, and multi-scale molecular simulation techniques in the research on phase separation. By comparing and analyzing the advantages and limitations of current related techniques in the research on phase separation, we make an outlook on the potential directions for future related research. In this review, we summarize the cutting-edge technologies related to plant phase separation, aiming to promote multidisciplinary integration and provide technical support for deciphering the mechanisms and functions of plant phase separation.

The role of alpha-ketoglutarate (AKG), a crucial metabolic intermediate, in enhancing the cryotolerance of mammalian sperm remains poorly understood. This study systematically investigated the effects of AKG on semen cryopreservation in dairy goats and elucidated its underlying mechanisms. Semen samples were collected from 10 healthy Saanen dairy goats, pooled after screening for motility (> 80%), and cryopreserved in liquid nitrogen vapor with extender supplemented with varying AKG concentrations (0, 10, 100, and1 000 μg/mL). Post-thaw assessments included sperm motility parameters, survival index, plasma and acrosome membrane integrity, antioxidant enzyme activities, ATP levels, and apoptosis rate. The optimal AKG concentration (100 μg/mL) was selected for subsequent analysis using non-targeted metabolomics to identify altered metabolites and pathways. In vitro fertilization and artificial insemination were conducted to comprehensively assess fertilizing capacity. Compared to the control, 100 μg/mL AKG most effectively enhanced post-thaw sperm quality, improving motility to 58.71%, and plasma and acrosome integrity to 61.56% and 62.49%, respectively. This concentration significantly elevated total antioxidant capacity, catalase, and superoxide dismutase activities, while reducing malondialdehyde and reactive oxygen species levels. Additionally, ATP content increased and the early apoptosis rate decreased. Non-targeted metabolomics revealed 397 significantly differential metabolites, predominantly enriched in 154 key pathways including the tricarboxylic acid cycle, pyruvate metabolism, glutathione metabolism, and the NF-κB signaling pathway. In vitro fertilization results showed that the addition of 100 μg/mL AKG significantly improved the cleavage rate of in vitro fertilized embryos but had no significant effect on the blastocyst rate. Additionally, the pregnancy rate from artificial insemination was significantly improved (34.13% vs. 21.97%), with a 12.16% increase. In conclusion, supplementing cryopreservation extender with 100 μg/mL AKG enhances post-thaw sperm function and fertility in dairy goats by modulating critical metabolic pathways related to energy metabolism and oxidative defense. This study provides a novel strategy and theoretical foundation for optimizing semen cryopreservation.

Wheat is one of the important food crops in China. Wheat sharp eyespot, caused by the infection of Rhizoctonia cerealis, is a destructive disease that seriously affects the yield and quality of wheat. Cultivating and popularizing disease-resistant varieties represents the most cost-effective and efficient approach for controlling wheat sharp eyespot. The resistance to wheat sharp eyespot is a quantitative trait regulated by multiple genes. This paper comprehensively reviews the advancements in genetic research on wheat resistance to sharp eyespot. It details the identification of wheat germplasm resources with resistance to sharp eyespot. A total of 51 reported disease-resistant QTLs are summarized, among which 33 exhibit a contribution rate exceeding 10% to phenotypic variation. Furthermore, this paper meticulously expounds on the research progress in the genes associated with wheat resistance to sharp eyespot and systematically analyzes the resistance mechanisms of various types of genes encoding transcription factors, protein kinases, key metabolic enzymes, epigenetic modification enzymes, and antimicrobial peptides. Finally, this paper sums up the research achievements in aspects such as the genome sequencing of R. cerealis and the interaction mechanism between R. cerealis and wheat. This review aspires to provide theoretical guidance for the genetic improvement of wheat resistance to sharp eyespot and the breeding of resistant varieties.

The genus Penicillium holds significant application value in industrial, biodegradation, and medical fields. However, the precise identification of its closely related species remains challenging. Analysis of whole-GEnome (AGE) is a novel approach for accurate species identification based on the screening and identification of species-specific target sequences from the whole genome. To address the identification challenges of closely related species within this genus, this study employed AGE to conduct bioinformatics analysis on 130 genomes from 82 closely related species of Penicillium, constructing species-specific target sequence libraries. After deduplication, the number of specific target sequences ranged from 2 562 to 557 355. To verify the applicability of these specific sequences, we selected seven representative species: Penicillium canescens, Penicillium oxalicum, Penicillium citrinum, Penicillium paneum, Penicillium roqueforti, Penicillium rubens, and Penicillium polonicum. One specific target sequence was screened for each species, and Sanger sequencing and CRISPR-Cas12a technology were employed to examine the reliability of the sequences. The results demonstrated that these seven specific sequences exhibited high specificity and applicability, effectively distinguishing P. canescens, P. citrinum, and P. rubens, which were challenging to be identified accurately. This confirmed the reliability of the specific target sequences. Additionally, these sequences were located in unannotated regions of the genome, showing excellent distinguishing ability. Furthermore, AGE enables the visual detection of the seven species, offering advantages for on-site testing. This study provides a reliable tool for resolving the challenges of identifying closely related species of Penicillium, offering valuable insights for the precise identification of other closely related fungal species.

Rhodotorula toruloides is an excellent oleaginous yeast species characterized by high lipid productivity, a broad substrate spectrum, and strong stress resistance. In addition, it has the suitability for high-density cultivation and the ability to synthesize carotenoids, which make it a potential chassis strain in the field of biotechnology. Signal peptides, as short peptide sequences guiding protein trafficking, have been widely employed in metabolic engineering and synthetic biology research. However, studies on signal peptides in R. toruloides remain limited. In this study, we investigated the organelle-targeting performance of three targeting sequences in R. toruloides, which direct proteins to the endoplasmic reticulum, mitochondria, and peroxisomes. After fusion of the targeting signal peptides with enhanced green fluorescent protein (eGFP), green fluorescence signals were observed in the corresponding organelles, confirming the utility of these signal peptides for protein localization. Furthermore, when hemoglobin VHb as the target protein was linked to the targeting sequences and eGFP, eGFP was still localized in the respective subcellular organelles, demonstrating that fusion with targeting signal peptides enabled the specific compartmentalization of target proteins. Finally, fatty acid photodecarboxylase (CvFAP) was expressed in R. toruloides with targeted localization, and the production of alkanes was detected in the engineered strains after fermentation under blue light. These results provide novel strain resources for the synthetic biology research on R. toruloides and expand its application in cell compartmentalization engineering.

Cytokinins, a class of plant hormones that can regulate the growth and development of plant cells, are closely related to crop yields and essential for promoting callus growth and differentiation. Cytokinin oxidases/dehydrogenases (CKXs) irreversibly catalyze the inactivation of cytokinins by side chain breakage and can dynamically regulate the cytokinin content in plants, thus participating in the photomorphogenesis, growth, and leaf senescence. However, the roles of OsCKXs in callus regeneration remain unclear. In this study, the plant tissue culture and cytokinin assay were employed to systematically investigate the changes in cytokinin content and regenerative capacity of callus from CKX family mutants in rice. We found that the mutation of OsCKX2 significantly increased the content of cytokinins and up-regulated the expression of regeneration-related genes WOX11 and LEC1 in the callus during regeneration, thereby increasing the regeneration rate. The mutation of OsCKX1 and OsCKX7 significantly reduced the content of cytokinins, induced severe browning, and down-regulated the expression of WOX11 and LEC1 in the callus during regeneration, thus reducing the regeneration rate. The treatment with cytokinins significantly mitigated the callus browning and improved the regeneration rate of osckx1 and osckx7 mutants, which indicated that cytokinin content was involved in callus browning. RT-qPCR results showed that the expression of OsCKX2 was high, while that of OsCKX1 and OsCKX7 was very low in the process of callus regeneration. These results indicate that OsCKX2, OsCKX1, and OsCKX7 are involved in the regulation of rice callus regeneration. The findings provide a theoretical basis for improving rice callus regeneration.

Trans-3-methyl-2-hexenoic acid (E3M2H) is a widely recognized as one of the key components leading to bromhidrosis. At present, the screening of E3M2H-degrading strains and the research on the molecular mechanism underpinning the efficient microbial degradation of E3M2H remain to be carried out. In this study, Rhodococcus qingshengii SDb-8 with high E3M2H degradation efficiency was isolated from forest soil and bath wastewater samples. Furthermore, the differential metabolic pathways of E3M2H and its structural analogues in the strain were analyzed by non-targeted metabolomics. R. qingshengii SDb-8 was found to raise the levels of substances such as 2,2-dimethylvaleric acid and 4-methylhexanoic acid during the oxidation of E3M2H by this strain. Furthermore, the expression levels of genes encoding diaminobutyric-2-oxoglutarate transaminase, L-2,4-diaminobutyric acid acetyltransferase, and L-carnosine synthase were significantly upregulated. This study not only provides a new microbial resource for the biodegradation of E3M2H but also lays an experimental foundation for elucidating the microbial degradation mechanism of E3M2H and provides a theoretical basis for developing a new strategy for the treatment of bromhidrosis based on microbial degradation.

The co-contamination of fluorene and heavy metals in industrial wastewater has become an environmental problem arousing wide concern. The presence of heavy metals impedes microbial degradation, making the identification of heavy metal-resistant degraders of fluorene a crucial research focus for bioremediation efforts. To address the current bottleneck that most reported degraders exhibit restricted activity and reduced degradation efficiency under heavy metal coexistence, this study selected two polycyclic aromatic hydrocarbon-degrading strains, Rhizobium sp. BX and Pseudomonas sp. DL, which demonstrated remarkable heavy metal tolerance. A synthetic mixed system was constructed from these two strains to investigate their fluorene degradation performance and mechanisms under heavy metal stress. Experimental findings indicated complete degradation of fluorene (25-50 mg/L) within 120 h. In addition, strains BX and DL exhibited tolerance to individual heavy metal ions [Cu²⁺, Zn²⁺, Pb²⁺, Cr(Ⅵ)] at 0-200 mg/L and combined heavy metal systems at 0-400 mg/L. A BX-DL mixed system was established, and its biodegradation conditions under heavy metal stress were optimized by response surface methodology. The optimal conditions (strain quantity ratio BX:DL=4:7, pH 8.59, 25.73 mg/L glucose, and inoculum volume of 8.91%) resulted in a degradation rate of 76.25% for 100 mg/L fluorene within 120 h. Whole-genome sequencing and metabolite profiling revealed complementary degradation pathways involved in salicylic acid and phthalic acid intermediates. This study provides potential microbial resources for the biological treatment of fluorene-containing wastewater under heavy metal stress.

Immunoglobulins are adaptive immune effector molecules unique to jawed vertebrates. Their remarkable diversity relies on mechanisms including V(D)J recombination, somatic hypermutation, and class switch recombination. These processes are orchestrated by a variety of key enzymes, such as recombination-activating gene proteins, terminal deoxynucleotidyl transferase (TdT), activation-induced cytidine deaminase, and DNA glycosylases. These enzymes not only play critical roles in the formation of antigen receptor diversity but have also been widely developed as useful tools for basic biological research. This review summarizes the molecular mechanisms underlying antigen receptor diversification in mammals, discusses recent progress in antibody screening and application, and highlights the latest advances in the use of TdT, cytidine deaminases, and DNA glycosylases in synthetic biology. These studies provide important theoretical support and new directions for fundamental research and technological innovation in biotechnology and medicine.

Patatin-like phospholipase domain-containing 3 (PNPLA3) I148M polymorphism (SNP rsID: rs738409; PNPLA3-I148M), a key genetic susceptibility factor for metabolic-associated fatty liver disease (MAFLD), is closely associated with disease progression. However, its mechanism in liver fibrosis remains to be elucidated. This study aimed to investigate the effects of PNPLA3-I148M overexpression on cholesterol metabolism, mitochondrial function, and fibrosis in the liver. We constructed the mouse models specifically overexpressing either the wild-type PNPLA3 (PNPLA3-WT) or PNPLA3-I148M in the liver. Liver fibrosis was induced via a high cholesterol-methionine and choline deficient (HC-MCD) diet. Our results showed that compared with the PNPLA3-WT group, mice overexpressing PNPLA3-I148M exhibited significantly elevated levels of total cholesterol, triglycerides, and free cholesterol, increased lipid droplet accumulation, and exacerbated steatosis and fibrosis in the liver. Mechanism studies revealed that PNPLA3-I148M interfered with cholesterol esterification and efflux by suppressing the expression of acyl-coenzyme A: cholesterol acyltransferase 1 (ACAT1) and ATP-binding cassette transporter G1 (ABCG1). This led to abnormal accumulation of free cholesterol in the liver, activated dynamin-related protein 1 (DRP1), and ultimately induced mitochondrial dysfunction and the expression of fibrosis-related genes. These findings provide important in vivo evidence and reveal a potential molecular mechanism by which PNPLA3-I148M promotes the development of liver fibrosis, laying the groundwork for developing precision therapeutic strategies targeting PNPLA3-I148M in MAFLD.