N⁴ -acetylcytidine (ac⁴C) modification is a chemical modification formed by the addition of an acetyl group to the nitrogen atom at position 4 of cytidine. It is widely distributed in various types of RNA molecules, including ribosomal RNA (rRNA) and transfer RNA (tRNA). In recent years, studies have revealed that ac⁴C also exists in messenger RNA (mRNA) as a novel mRNA modification with relatively low abundance. The ac⁴C modification is mainly catalyzed by the RNA acetyltransferase N-acetyltransferase 10 (NAT10) and plays vital regulatory roles in key biological processes such as ribosome biogenesis, codon recognition, and mRNA translation. In plant mRNAs, ac⁴C is predominantly enriched in the 5' untranslated region (5' UTR) near the start codon, and this modification can significantly enhance mRNA stability and translation efficiency. In model plants and crops such as Arabidopsis thaliana and rice, ac⁴C modification participates in regulating multiple biological processes including leaf development, photosynthetic efficiency, fruit ripening, and biotic stress responses. Currently, research on ac⁴C modification is limited by detection technologies. Existing ac⁴C detection methods (such as acRIP-seq and ac⁴C-seq) yield inconsistent results, leading to ongoing controversies regarding the abundance and function of ac⁴C on mRNA. This review summarizes the distribution characteristics of ac⁴C and the functions of the RNA acetyltransferase NAT10, compares the advantages and disadvantages of different ac⁴C detection techniques, discusses the physiological functions and molecular mechanisms of ac⁴C in plants, and prospects the challenges and future directions in this field. This review not only provides a theoretical basis for further dissecting the dynamic regulatory network and molecular mechanisms of ac⁴C, but also offers references for promoting crop genetic improvement based on RNA modifications.

Weeds are a major limiting factor in rice production, as they not only cause significant reductions in rice yield and quality but also escalate the costs of field management. The development of herbicide-resistant rice varieties is therefore a core strategy for efficient weed control in rice cultivation, as it can effectively extend the application scope and period of herbicides. Gene editing technology, endowed with efficient, precise, and rapid targeted modification capabilities, has overcome the inherent bottlenecks of traditional breeding approaches-including long breeding cycles, poor directional selectivity, and heavy reliance on elite germplasm resources-thus providing a revolutionary technical tool for the development of herbicide-resistant crops. This paper systematically reviews the latest research progress in the application of gene editing technology to herbicide-resistant rice breeding and briefly elaborates on the core working principles of the three most widely adopted gene editing systems to date: CRISPR/Cas-mediated NHEJ/HDR, base editing, and prime editing. It focuses on the applications of these three systems in the genetic improvement of key herbicide target genes in rice, including ALS, ACCase, EPSPS, GS, HPPD, and TubA2. Additionally, this study summarizes the mutation types induced by gene editing, along with the corresponding types and levels of herbicide resistance achieved in rice following the modification of these target genes. Finally, in light of the current challenges confronting the development and breeding of herbicide-resistant rice, prospects are proposed for the further optimization and advancement of gene editing technologies, the mining of novel functional genes, and the enhancement of environmental safety and sustainable development in this field. This review aims to provide a comprehensive reference for subsequent research and practical applications of new herbicide-resistant rice varieties.

Paclitaxel is a potent tetracyclic diterpenoid anticancer agent widely used in the clinical treatment of various malignancies, including ovarian and breast cancers. Its conventional production relies on extraction from Taxus spp. (yew trees), which is constrained by resource scarcity and ecological risks. Plant endophytic fungi are promising alternative sources for paclitaxel biosynthesis, offering potential for sustainable production and industrial-scale application. This paper systematically reviews the research progress on paclitaxel production by plant endophytic fungi. In terms of isolation sources, paclitaxel-producing endophytic fungi have been isolated from diverse host plants, including gymnosperms (e.g., Taxus), angiosperms (e.g., Corylus avellana), and herbaceous plants (e.g., Artemisia argyi). Regarding microbial classification, the reported paclitaxel-producing endophytic fungi span at least 15 fungal genera, including Alternaria, Aspergillus, and Penicillium. Wild-type strains show remarkable differences in paclitaxel biosynthetic capacity, with yields ranging from micrograms per liter to milligrams per liter. Due to the heterogeneity of culture conditions and the lack of unified quantitative detection standards, no dominant high-yield strains have been clearly identified to date. At the genetic level, homologous sequences of several key genes involved in the plant paclitaxel biosynthetic pathway have been identified in paclitaxel-producing endophytic fungi, whereas the complete paclitaxel biosynthetic pathway in endophytic fungi has not been fully elucidated. Current efforts mainly focus on optimizing endophytic fungal chassis cells through strain screening, mutagenesis breeding, and gene editing. In parallel, strategies such as fermentation process optimization, metabolic regulation, and co-culture have also shown significant potential for improving paclitaxel yields in endophytic fungi. Against the backdrop of the rapidly developing field of synthetic biology, this paper proposes that future research should strengthen integrated multi-omics analysis of endophytic fungi, along with the development of synthetic biology tools and metabolic engineering approaches, so as to provide a theoretical basis and technical support for the industrial production of paclitaxel using fungi as chassis cells.

Salt stress, caused by excessive accumulation of Na⁺ and Cl⁻ ions in soil, is a major abiotic stress that severely disrupts ionic homeostasis and water uptake in plants and induces excessive production of reactive oxygen species (ROS), thereby inhibiting growth and reducing crop yield and quality. With the rapid development of protected cultivation, secondary soil salinization has become increasingly severe. Cucumber (Cucumis sativus L.) is highly sensitive to salt stress, making the screening of salt-tolerant germplasm and elucidation of its molecular regulatory mechanisms a critical research focus for improving production efficiency. At present, salt-tolerant cucumber germplasm is mainly derived from cultivated varieties, landraces, and wild relatives. Their tolerance to salt is commonly evaluated using growth, physiological, and biochemical traits in combination with comprehensive indices such as salt tolerance indices and membership function analysis. Accumulating evidence indicates that tolerance in cucumbers to salt is a complex quantitative trait governed by coordinated regulation of multiple genes and relies on the integration of ionic homeostasis, osmotic adjustment, and antioxidant defense. Key processes include Na⁺ exclusion and compartmentalization, maintenance of K⁺ homeostasis, accumulation of osmotic adjustment substances, and ROS scavenging by antioxidant systems, with associated genes playing important roles in signal transduction and metabolic regulation. This review summarizes salt-tolerant germplasm resources and screening and evaluation methods in cucumber, outlines the major molecular regulatory mechanisms underlying salt tolerance, and, in light of recent advances in multi-omics and high-throughput phenotyping technologies, discusses future directions for molecular breeding of salt-tolerant cucumber, providing a theoretical basis and scientific reference for the development of highly salt-tolerant cultivars.

Genetic pleiotropy refers to the biological phenomenon in which a single gene or genetic variation simultaneously influences multiple phenotypic traits or diseases. With the rapid development of omics technology, pleiotropy research has achieved significant advances in revealing the genetic basis and molecular mechanisms of human complex traits and diseases, and a large number of pleiotropic genes and functional genetic variant sites have been identified. Important economical traits in livestock and poultry are predominantly complex in nature and commonly exhibit both phenotypic and genetic correlations, and genetic pleiotropy is one of the important mechanisms underlying this phenomenon. However, current studies on pleiotropy related to important economic traits in livestock and poultry remain relatively limited, primarily due to constraints in research strategies and technical approaches. By adapting successful methodologies from studies of human complex traits and diseases and applying them to livestock and poultry, it is conducive to deeply understanding the shared genetic architecture among correlated traits, clarifying the nature of these correlations (synergistic or antagonistic), thereby ultimately providing a robust theoretical basis for designing scientifically sound, rational, and efficient breeding programs. This review systematically summarizes the historical development, molecular mechanisms, commonly used study methods of genetic pleiotropy, and advances in the analysis of pleiotropy in important economic traits of livestock and poultry. It aims to provide references for further elucidation of the genetic basis of these traits, facilitating the discovery of pleiotropic genes and related molecular markers, and promoting genetic improvement in livestock and poultry.

Silage is a key technology for preserving green feed in animal husbandry, and its quality directly determines the production performance of ruminants. As silage additives, Bacillus species can significantly improve silage fermentation quality by regulating the microbial community structure, thereby promoting the accumulation of organic acids and inhibiting the reproduction of spoilage bacteria. Bacillus subtilis (BS) and Bacillus licheniformis (BL) are currently the two most widely used Bacillus species in the silage field, but their application effects show significant differences. This article systematically reviews the application characteristics of BS and BL in silage production, conducts a comparative analysis of the aspects of biological characteristics, mechanisms of action, fermentation effects, raw material adaptability and practical applications, and integrates the comparative study data of the two species under the same experimental conditions. The results show that BS has prominent advantages in rapidly reducing the pH value of the silage system, degrading non-structural carbohydrates, and secreting high-activity protease; while BL performs better in improving silage aerobic stability, inhibiting mycotoxins, and degrading structural carbohydrates. The synergistic application of the two species can achieve functional complementarity, significantly improve silage quality and reduce production costs. This review can provide a solid theoretical basis for the scientific selection and efficient application of silage additives.

Objective This study aimed to systematically evaluate the potential of the CRISPR-Cas12a (Cpf1) system mediating site-specific gene knock-in in porcine cells, thereby providing an experimental basis and theoretical reference for its application in precise porcine genome engineering. Method Expression vectors for Acidaminococcus sp. Cas12a (AsCas12a) and Lachnospiraceae bacterium Cas12a (LbCas12a) were constructed. Specific crRNAs were designed to target the well-characterized porcine “safe harbor” loci, Rosa26 and AAVS1, to compare the cleavage efficiencies of the two Cas12a variants. Result At the AAVS1 locus, the editing efficiencies of two crRNAs for AsCas12a were 2.6% and 1.2%, respectively, whereas those for LbCas12a were 6.5% and 2.5%. At the Rosa26 locus, the editing efficiencies for AsCas12a and LbCas12a were 5.0% and 12.7%, respectively, suggesting that LbCas12a possessed relatively higher editing activity in porcine cells. Based on these findings, knock-in donors were developed to utilize four distinct DNA repair pathways: Homology-directed repair (HDR), homology-mediated end joining (HMEJ), microhomology-mediated end joining (MMEJ), and non-homologous end joining (NHEJ), and their integration efficiencies were evaluated. The results revealed that 4 repair pathways all mediated the target genes to have site-targeted integration at both Rosa26 and AAVS1 loci. Specifically, the efficiency by HDR achieved 35.0%, 25.0% by HMEJ, and 15.0% by both NHEJ and MMEJ. Conclusion Four site-specific integration strategies are successfully developed in porcine cells using the Cas12a gene-editing system, followed by a systematic evaluation of their efficiencies. These findings provide essential experimental evidence and references for expanding the application of the Cas12a system in porcine genetic engineering.

Objective To address the drawbacks in field applications, such as complex operation and long detection cycles, for porcine reproductive and respiratory syndrome virus type Ⅱ (PRRSV-2), this study aimed to develop a rapid and convenient detection method for disease surveillance and epidemic control. Method Primers and CRISPR RNAs (crRNAs) were designed based on conserved regions of the PRRSV-2 genome. A one-pot detection system integrating recombinase-aided amplification (RAA) and CRISPR/EsCas13d was established. Through systematic optimization of reaction component concentration and reaction conditions, RAA amplification, T7 transcription, and CRISPR/EsCas13d cleavage were sequentially completed in a single reaction tube, eliminating tube-opening steps during the assay. Detection results were obtained using a fluorescence instrument or by direct naked-eye visualization of in-tube fluorescence signals. Sensitivity, specificity, and clinical performance were evaluated and compared with reverse transcription quantitative polymerase chain reaction (RT-qPCR). Result The one-pot RAA-CRISPR/EsCas13d detection platform enabled nucleic acid detection of PRRSV-2 within 60 min. Sensitivity analysis showed a detection limit of 8 copies/μL with fluorescence readout and 50 copies/μL with naked-eye visualization. Specificity analysis demonstrated no cross-reactivity with common porcine viruses, including classical swine fever virus and porcine epidemic diarrhea virus. For 56 clinical samples, detection results were fully consistent with those obtained by RT-qPCR. Conclusion The one-pot RAA-CRISPR/EsCas13d detection platform enables sensitive detection of PRRSV-2 within 60 min. This platform supports fluorescence-based readout and naked-eye visualization, and provides a simple and rapid molecular diagnostic method for PRRSV-2 detection and disease surveillance.

Objective The mSA-biotin system was used to recruit donor DNA to target sites, enhancing the editing efficiency of exogenous genes in homology-directed repair (HDR) via RNP-mediated CRISPR/Cas9 and the efficiency in scarless gene editing of filamentous fungi. Method By utilizing the binding ability of streptavidin (mSA) and biotin, mSA was fused with Cas9 for expression, and recombinant protein Cas9-mSA was obtained. Subsequently, donor DNA was amplified by PCR using a 5′-biotinylated primer, yielding biotinylated donor DNA (Bio-Donor DNA). Assembling Cas9-mSA and sgRNA into active RNP-mSA complexes in vitro, and they were co-transformed into Aspergillus tubingensis F316 strain with Bio-Donor DNA. While the spore pigment-related gene fwnA in its genome was knocked out, and the green fluorescent protein EGFP was simultaneously integrated into the fwnA gene site through HDR. Then, the positive transformants were screened through spore color and PCR detection quickly, and HDR efficiency was statistically calculated. Result The use of Cas9-mSA with Bio-donor DNA makes the donor DNA directly enriched toward gene editing sites, and thus HDR efficiency increased by 56.53%, which promoted the application of RNP method in gene editing of filamentous fungi. Conclusion The combined use of Cas9-mSA and Bio-Donor DNA effectively enhances the HDR efficiency of the RNP method in filamentous fungi.

Objective Analysis of quantitative trait loci (QTLs) associated with rice leaf wilt degree, providing a reference for screening drought-tolerance candidate genes in rice. Method Using a panel of 300 rice germplasm resources, the leaf rolling index (LRI) and leaf wilt degree (LWD) were investigated under drought stress. Genome-wide association study (GWAS) was employed to map drought tolerance-related quantitative trait loci (QTLs). Result Phenotypic data for LRI and LWD followed a normal distribution. Four QTLs associated with LWD were identified via GWAS. Among these, qLWD4-1 was consistently detected across both investigation periods. Based on haplotype analysis and gene function annotation, four candidate genes for drought tolerance (LOC_Os04g54360, LOC_Os04g54474, LOC_Os04g54560, and LOC_Os04g54570) were screened within the qLWD4-1 locus region. Conclusion This study identified a stably expressed QTL (qLWD4-1) associated with leaf wilt degree across two investigation periods via GWAS, and four drought-resistant candidate genes were determined within this locus.

Objective This study aimed to systematically elucidate the genetic basis of the novel Japonica rice variety Jiru Dao 1, evaluate its production potential and risks, and formulate precise molecular design breeding strategies. Method Agronomic traits, grain quality, and disease resistance data from the Shandong Provincial Regional Trials and Production Trials (2019-2021) were integrated. Whole-genome resequencing of Jiru Dao 1 was performed using the Illumina HiSeq platform. Genotyping of 319 important agronomic trait-associated quantitative trait nucleotides (QTNs) was conducted based on the RiceNavi system, followed by genotype-phenotype association analysis. Result The average yield of Jiru Dao 1 was 10 437.0 kg/hm², 8.1% higher than the control. Its grain quality met the Level 3 standard of the national “High-Quality Paddy” (GB/T 17891-1999) specification, and it was recognized as a superior-tasting variety by Jiangsu province in 2023. Genotype analysis revealed that its high yield and lodging resistance originated from the pyramiding of favorable alleles such as sd1 and Ghd8; its excellent tasting quality was controlled by key genes including Waxy (low amylose content) and ALK (low gelatinization temperature). For disease resistance, it carried multiple blast resistance genes (pi35, Pita, Pib, Pid2, Pid3, and Pi21) and genes (Xa3, xa25) resisting to bacterial blight. Both OsCd1 and OsNRAMP5 were superior “non-cadmium-accumulating” alleles. Meanwhile, Gn1a and NOG1 were identified as “to-be-improved” alleles limiting yield potential. Chalk5 was a “chalkiness-increasing” allele affecting appearance quality; and NRT1.1B, OsNR2, and PSTOL1 were mostly “neutral” alleles, constraining nutrient use efficiency. Conclusion Jiru Dao 1 represents a successful example of traditional breeding by pyramiding multiple favorable alleles for high yield, good quality, multiple disease resistance, and low cadmium accumulation. However, its yield potential, appearance stability, and disease resistance breadth are limited by inferior alleles such as Gn1a and Chalk5 and the absence of broad-spectrum resistance genes. Whole-genome dissection precisely identified these genetic strengths and weaknesses, providing a scientific basis to overcome the empirical limitations of conventional breeding. Therefore, a precision breeding strategy is proposed: using Jiru Dao 1 as the recipient, key favorable alleles (Gn1a, Chalk5, Pi2/Pi9,and Xa23) are introduced via marker-assisted selection, while whole-genome background selection preserves its elite genetic background, enabling targeted variety improvement.

Objective The LAX1-LAX2 complex is a key module regulating axillary meristem development in the rice panicle. The bHLH (basic helix-loop-helix) transcription factor OsbHLH069 interacts with LAX1, and its gain-of-function mutant nsp1-D (no spikelet 1-dominant) exhibits a sparse panicle phenotype. This study aimed to elucidate the molecular interaction model among these three proteins and clarify the mechanism by which they regulate panicle branching development. Methods The nuclear localization signal and transcriptional activation domain of OsbHLH069 were analyzed using rice protoplasts and yeast systems. The interaction between OsbHLH069 and LAX2 was investigated through yeast two-hybrid assays, bimolecular fluorescence complementation, and in vitro pull-down assays. The lax2-4 nsp1-D double mutant was generated by field crossing, and genetic interactions were analyzed by quantitative real-time PCR. The interaction regions among the three proteins were mapped using truncation mutants and yeast two-hybrid assays. The interaction model was further explored via luciferase complementation assays. Results OsbHLH069 is a typical nuclear-localized transcriptional activator, with its nuclear localization signal located at amino acids 266–285 and its transcriptional activation domain at the N-terminal region (amino acids 1–120). OsbHLH069 directly interacted with LAX2 both in vivo and in vitro. OsbHLH069 interacted with LAX1 through its bHLH domain and with LAX2 through its C-terminal region; these interaction regions overlapped with the LAX1-LAX2 interaction interface. OsbHLH069 competitively inhibited the stability of the LAX1-LAX2 complex. The lax2-4 nsp1-D double mutant showed more severe defects in branch number and spikelets per panicle than either single mutant, resembling the previously reported phenotype of the lax1 nsp1-D double mutant. Conclusion A phased regulatory model is proposed: during early panicle axillary meristem development, the LAX1-LAX2 complex promotes axillary meristem initiation; as development progresses, OsbHLH069 achieves fine-tuned negative regulation of the panicle axillary meristem by competitively disassembling the LAX1-LAX2 complex.

Objective To systematically identify the members of the wheat homeodomain-leucine zipper (HD-Zip) gene family, analyze their expression patterns across different tissues and developmental stages, and investigate their roles in response to Fusarium graminearum, thereby providing a theoretical basis for functional characterization of the wheat HD-Zip gene family. Method Using Arabidopsis HD-Zip protein sequences as queries, the members of the wheat HD-Zip gene family were identified from the wheat genome database. Protein physicochemical properties, phylogenetic tree construction, chromosome mapping and gene structure analysis were analyzed. Furthermore, transcriptome data and RT-qPCR were employed to assess the expression profiles of these genes in various tissues and under F. graminearum infection. Result A total of 85 wheat HD-Zip genes were identified, all of which encoded hydrophilic proteins, and were predicted to be unstable. Subcellular localization indicated that most HD-Zip proteins were localized in the nucleus and chloroplast. Phylogenetic and structural analyses revealed that these genes were grouped into four evolutionary clades, with members of the same clade showing high similarity in gene structure and conserved motifs, and were unevenly distributed across 22 chromosomes. Promoter analysis indicated the widespread presence of cis-acting elements associated with abscisic acid (ABA), gibberellin (GA), methyl jasmonate (MeJA), salicylic acid (SA), and auxin (IAA), as well as elements involved in stress responses, root specificity, and endosperm regulation. Expression analysis showed that 65 genes were expressed in the spikes, leaves, shoots, roots, and grains. Notably, under F. graminearum infection, genes such as TaHDZ1.1-6B, TaHDZ1.1-5A, TaHDZ1.1-5B, and TaHDZ1.1-5D were significantly upregulated at multiple time points, exhibiting consistent inducible expression patterns in both resistant and susceptible cultivars. Conclusion The wheat HD-Zip gene family presents strong evolutionary conservation and is likely involved in both organ development and responses to biotic and abiotic stresses through differential expression. Among them, TaHDZ1.1-6B, TaHDZ1.1-5A and TaHDZ1.1-5B may play crucial roles in wheat defense against F. graminearum.

Objective Pseudo-response regulators (PRRs) play crucial roles in regulating circadian rhythms and flowering time in plants. This study aimed to systematically identify the PRR gene family in broomcorn millet (Panicum miliaceum L.) at the whole-genome level, providing a theoretical basis for understanding their biological functions. Method Bioinformatics methods were employed to analyze the physicochemical properties, conserved motifs, gene structures, collinearity, and phylogenetic relationships of the PRR gene family in broomcorn millet. The expression patterns of PRR genes under different photoperiod treatments were investigated using real-time quantitative PCR (RT-qPCR). Result A total of seven PRR genes were identified from the broomcorn millet genome, designated as PmPRR1 to PmPRR7, which were unevenly distributed across five chromosomes. Phylogenetic analysis revealed that the PmPRR genes belong to Group 1 to Group 3 subfamilies. Analysis of conserved protein motifs showed that six genes contained Motif1, Motif2, and Motif3, indicating high conservation among them. Promoter cis-acting element analysis suggested that PmPRR genes are associated with light response, plant growth and development, stress responses, and hormone regulation. RT-qPCR results demonstrated significant differences in the expressions of PmPRR genes under different photoperiod treatments (short-day, 8 h; intermediate-day, 14 h; long-day, 16 h). Conclusion This study identified seven PmPRR genes. The expression patterns of genes from different subfamilies vary under different photoperiods, indicating that PmPRRs play an important regulatory role in the response of broomcorn millet to photoperiod changes.

Objective NAC (NAM/ATAF1/2/CUC2) transcription factors are a crucial class of regulatory proteins in plants, extensively involved in growth, development, and responses to biotic and abiotic stresses. This study aimed to clone the HvNAC53 gene from hulless barley (Hordeum vulgare var. nudum), analyze its sequence characteristics and expression patterns, and functionally characterize it, thereby providing a theoretical basis for stress-tolerant molecular breeding in hulless barley. Methods The HvNAC53 gene was cloned from the hulless barley cultivar ‘Kunlun 14’. Bioinformatics analysis and RT-qPCR were used to analyze gene expression and protein characteristics. The role of HvNAC53 in response to low-temperature and drought stress was investigated through heterologous overexpression in Arabidopsis thaliana. Results The open reading frame of HvNAC53 is 1 550 bp in length, encoding a protein of 305 amino acids. The protein lacks transmembrane domains and signal peptides and is classified as a hydrophilic, unstable protein. Multiple cis-acting elements responsive to light, hormones, and stress were identified in the promoter region of HvNAC53. Phylogenetic analysis indicated that HvNAC53 is most closely related to TeNAC53 from Thinopyrum elongatum (tall wheatgrass). RT-qPCR results revealed high expression levels of HvNAC53 in roots and leaves compared to other tissues, and its expression was significantly induced by drought and cold stress. Subcellular localization demonstrated that HvNAC53 is localized in the nucleus. Furthermore, Arabidopsis thaliana lines heterologously overexpressing HvNAC53 exhibited significantly higher cold and drought tolerance than wild-type plants. Conclusion HvNAC53 positively regulates low-temperature and drought stress responses in hulless barley.

Objective Type 2C protein phosphatases (PP2Cs) are core components in the abscisic acid (ABA) signal transduction pathway and play crucial roles in plant responses to abiotic stresses. This study aimed to clone PP2C family genes from apple and investigate their roles in ABA signaling and drought stress responses, providing a theoretical basis and genetic resources for apple stress-resistant breeding. Method The MdPP2C24 gene was cloned from ‘Gala’ apple. Phylogenetic tree and protein structure analyses were performed to identify conserved domains and homology. Quantitative real-time PCR (RT-qPCR) was used to detect expression patterns, transient expression in onion inner epidermis was employed to observe protein localization, and MdPP2C24-overexpressing Arabidopsis thaliana plants were generated via floral dip method to analyze ABA sensitivity and drought tolerance. Result The open reading frame of MdPP2C24 was 1 218 bp, encoding 405 amino acids with a molecular weight of 44.29 kD and the pI was 6.63. Phylogenetic and protein structure analyses revealed that MdPP2C24 was closely related to PbPP2C24 from pear (Pyrus × bretschneideri) and contained one conserved PP2C domain. RT-qPCR showed that MdPP2C24 was expressed in all apple tissues, with the highest expression in the roots, and its expression was significantly upregulated by ABA and mannitol treatments. Subcellular localization indicated that MdPP2C24 was a nuclear-cytoplasmic co-localized protein. Furthermore, heterologous overexpression of MdPP2C24 in Arabidopsis significantly reduced ABA sensitivity and drought tolerance, while inhibiting the expression of stress-responsive marker genes. Conclusion The apple protein phosphatase gene MdPP2C24 reduces plant drought tolerance by negatively regulating ABA sensitivity and suppressing the expression of stress-responsive marker genes.

Objective Fatty acyl-ACP thioesterase (FAT) plays a key role in plant oil synthesis. This study aimed to characterize FAT family genes in yellow nutsedge (Cyperus esculentus L.) and to provide a theoretical basis and technical support for breeding high-oil cultivars and improving lipid metabolism. Method Based on the whole-genome data of yellow nutsedge, the CeFAT gene family was systematically identified and subjected to sequence and phylogenetic analyses. The subcellular localization of CeFATA was determined, and the function of CeFATA was evaluated using yeast and tobacco genetic transformation systems. Result Three CeFAT genes, CeFATA, CeFATB1, and CeFATB2, were identified. They showed similar gene structures and physicochemical properties, and were located on three different chromosomes. Phylogenetic analysis indicated that CeFATA was most closely related to OsFATA from rice, whereas CeFATB1 and CeFATB2 were closely related to GmFATB from soybean. RT-qPCR analysis showed that CeFAT genes were highly expressed at five developmental stages of the tuber, with CeFATA showing the highest expression level. Subcellular localization confirmed that CeFATA was located in chloroplasts. Overexpression of CeFATA increased total lipid content in Saccharomyces cerevisiae by 3.1% and exhibited substrate preference for C18:1. In transgenic tobacco, CeFATA overexpression increased total fatty acid contents in leaves and seeds by 4.74% and 8.4%, accompanied by a significant decrease in starch content, a decrease soluble sugar content in leaves, an increase soluble sugar content in seeds, and no significant change in protein content. Conclusion CeFATA increases total lipid and oleic acid contents in yeast and tobacco, promotes the export of oleic acid from plastids, and redirects carbon flux from carbohydrate biosynthesis toward oil biosynthesis.

Objective To elucidate the chloroplast genome characteristics of Actinidia rubricaulis var. coriacea and investigate its phylogenetic relationship within the genus Actinidia, providing a genetic basis for species identification, genetic diversity analysis, and resource conservation of Actinidia species. Method The complete chloroplast genome of A. rubricaulis var. coriacea was sequenced using the Illumina NovaSeq platform, assembled and annotated with bioinformatics software, and compared with chloroplast genome sequences of representative Actinidia species downloaded from GenBank. Result The chloroplast genome of A. rubricaulis var. coriacea is 156 547 bp in length, with a GC content of 37.24%. It is composed of a large single-copy region (88 649 bp), a small single-copy region (20 484 bp), and two inverted repeat regions (23 707 bp each), exhibiting a typical quadripartite structure. For this species, the chloroplast genome contains a total of 133 annotated genes, including 85 protein-coding genes, 39 transfer RNA (tRNA) genes, and 8 ribosomal RNA (rRNA) genes. Among these, 21 genes contain introns. Codon usage analysis revealed that among the 30 preferred codons with a relative synonymous codon usage value >1, 28 end with A or U. Repeat sequence analysis detected 312 long repeat sequences and 207 simple sequence repeats, with 128 single nucleotide repeats, accounting for 61.84%. Phylogenetic analysis showed that Actinidia rubricaulis var. coriacea is most closely related to Actinidia hubeiensis, with bootstrap support of 100%. Conclusion The chloroplast genome of A. rubricaulis var. coriacea is relatively conserved in structural characteristics and gene composition, but shows some variation in genomic boundaries and nucleotide polymorphism compared with other species in the same genus. It exhibits a higher preference for codons ending with A or U. Actinidia rubricaulis var. coriacea is closely related to Actinidia hubeiensis and highlights the need for further refinement of the taxonomic system within the genus Actinidia.

Objective This study aimed to investigate the response of the VvTPS10 gene to salt stress in grapevine (Vitis vinifera), providing theoretical insights for the study of salt tolerance mechanism in grapevine. Method After subjecting potted grape seedlings to salt stress treatment, the expression pattern of VvTPS10 was detected by RT-qPCR. Phenotypic observations of VvTPS10-overexpressing Arabidopsis thaliana and grape calli were performed after salt stress treatment, followed by the determination of physiological and biochemical indices and the analysis of stress-resistant gene expression. Result The expression of the VvTPS10 gene in grapevines was significantly up-regulated under salt stress. Under the treatments of 150 mmol/L NaCl and 200 mmol/L NaCl, the germination rates of VvTPS10-overexpressing A. thaliana were 18.04% and 50.57% higher than those of the wild-type, respectively. Under salt stress, the relative electrical conductivity, MDA content and superoxide anion content of VvTPS10-overexpressing A. thaliana were significantly lower than those of the wild-type, whereas the chlorophyll content, and the activities of SOD, POD and CAT were significantly higher than those of the wild-type. RT-qPCR analysis showed that under salt stress, the expression levels of stress-resistant genes including COR15A、COR47、RD29B、SOS1、NHX1 and DREB2A in VvTPS10-overexpressing plants were significantly higher than those in the wild-type. Meanwhile, we found that overexpression of VvTPS10 not only resulted in an early bolting and flowering in A. thaliana but also responded to exogenous sucrose treatment, as evidenced by a significant increase in anthocyanin accumulation. Finally, under salt stress, VvTPS10-overexpressing grape calli exhibited better growth status, higher fresh weight and lower relative electrical conductivity. Conclusion The VvTPS10 gene is induced by salt stress, and it enhances the salt tolerance of A. thaliana by maintaining cell membrane stability, improving antioxidant enzyme activities and promoting the expression of stress-resistant genes. In addition, this gene promotes early flowering of A. thaliana and increases anthocyanin accumulation under sucrose treatment. Similarly, overexpression of VvTPS10 enhanced salt tolerance in grape callus.

Objective Terpenoids play important roles in plant growth, development, and stress defense. This study aims to investigate the biological function of the tobacco sesquiterpene synthase gene NtEAS (Nicotiana tabacum5-epi-aristolochene synthase), and to provide a theoretical basis for revealing the terpene-mediated mechanisms of resistance and defense to disease and promoting molecular breeding in tobacco. Method Bioinformatics methods were employed to analyze the characteristics, conserved domains, cis-acting elements, and phylogenetic relationships of the NtEAS protein. RT-qPCR was used to detect the expression patterns of NtEAS in different tissues and under pathogen stress. Subcellular localization technology was applied to determine the distribution of NtEAS protein within cells. The NtEAS protein was expressed in vitro and purified using a prokaryotic expression system, and its enzymatic catalytic products were identified by GC-MS. The mycelial growth rate method and filter paper diffusion method were employed to evaluate the in vitro inhibitory activity of its catalytic products against tobacco bacterial wilt and black shank pathogens, in vitro prokaryotic expression and bacterial inhibitory activity analysis. Result The NtEAS has an open reading frame with a total length of 1647 bp, encoding 548 amino acids. Its encoded protein was located in the nucleus and cytoplasm and contains typical terpene synthase conserved domains “DDXXD” and “NST/DTE”. NtEAS was mainly highly expressed in tobacco roots and strongly induced by Ralstonia solanacearum and Phytophthora nicotianae. NtEAS can catalyze the formation of sesquiterpenoid aristolochene with farnesyl pyrophosphate as a substrate. The catalytic product exhibits significant inhibitory activity against both R. solanacearum and P. nicotianae, and this inhibitory effect is positively correlated with its concentrations. Conclusion NtEAS gene and its catalytic product may be involved in the chemical immune defense process of tobacco against bacterial wilt and black shank.

Objective To clone the NtPLA2A gene from tobacco (Nicotiana tabacum) and explore the molecular mechanism by which it mediates hormone signaling pathways to enhance tobacco resistance to bacterial wilt. Method This study cloned the NtPLA2A gene from tobacco, conducted bioinformatic analysis and expression profiling under Ralstonia solanacearum infection, and investigated the role of NtPLA2A in disease resistance using virus-induced gene silencing (VIGS) in N. benthamiana. Result The NtPLA2A coding sequence is 1 284 bp, encoding 428 amino acids. The protein has a molecular weight of 46.57 kD and a theoretical isoelectric point of 8.74, and is a stable hydrophilic protein. Bioinformatic analysis revealed that NtPLA2A contains a typical Patatin-like phospholipase domain and shares high amino acid sequence similarity with PLA proteins from other species. Promoter analysis indicated that the promoter region of NtPLA2A contains environmental factors such as light, low temperature, and hormones. Following R. solanacearum infection, NtPLA2A was significantly upregulated in both resistant and susceptible tobacco materials to varying degrees. Additionally, treatments with salicylic acid, jasmonic acid, and ethylene also induced its expression. VIGS results demonstrated that silencing NtPLA2A compromised resistance to R. solanacearum in N. benthamiana. Conclusion This study clarified the sequence, structure, and expression characteristics of tobacco NtPLA2A gene, which is induced by R. solanacearum and SA/JA/ET; its silencing significantly impairs N. benthamiana resistance to bacterial wilt.

Objective Glucose-6-phosphate dehydrogenase (G6PDH) plays a crucial role in plant growth, development, and stress responses. This study aimed to clone the G6PDH genes from safflower (Carthamus tinctorius L.), analyze their sequences and expression patterns, and investigate their involvement in abiotic stress responses. These findings will provide valuable candidate genes for stress-resistant safflower breeding. Method Based on the genome data of safflower, G6PDH genes were identified and cloned, followed by bioinformatic analysis. Using RT-qPCR, the tissue-specific expression patterns of CtG6PDH genes and their responses to hormones (MeJA, ABA, and SA) and drought, salt and alkali stress were analyzed. Additionally, the activities of glucose-6-phosphate dehydrogenase (G6PDH) under these abiotic stresses were determined by ultraviolet spectrophotometry. Result Two G6PDH family members (CtG6PDH1 and CtG6PDH2) were identified and cloned from the safflower genome, located on chromosomes 1 and 3, respectively. Subcellular localization and phylogenetic analysis revealed that CtG6PDH1 is a cytosolic isoform localized in the cytoplasm, while CtG6PDH2 belongs to the plastidial P1-type and is located in chloroplasts. Their promoter regions contain multiple cis-acting elements associated with light response, phytohormone signaling, and stress responses. Tissue expression analysis showed that CtG6PDH1 exhibits higher expression in roots, while CtG6PDH2 demonstrated the highest expression in leaves. Under drought and saline-alkaline stress, the cytosolic CtG6PDH1 gene was markedly upregulated, while CtG6PDH2 expression exhibited induction by multiple phytohormones. Further enzymatic assays confirmed that cytosolic G6PDH activity was significantly enhanced under both drought and saline-alkali stress conditions. Conclusion Two G6PDH genes were identified and cloned from safflower, exhibiting distinct subcellular localizations (cytosol and chloroplast). Promoter analysis revealed putative cis-regulatory elements associated with growth, hormone responses, and stress adaptation. RT-qPCR and enzyme activity analyses demonstrated that the cytosolic CtG6PDH1 plays a major role in safflower’s response to abiotic stresses, including drought, salt, and alkali.

Objective To investigate whether a formate dehydrogenase (FDH) gene RcFDH2 of Racomitrium canescens possesses drought- and high-temperature-tolerant functions, and to provide theoretical support for revealing the drought- and high-temperature-tolerant mechanisms of R. canescens and for cultivating resistant crops. Method Real-time fluorescence quantitative PCR technology was used for the detection of the expression variations of RcFDH2 during the rehydration process of desiccated R. canescens. The coding sequence of RcFDH2 was successfully cloned. RcFDH2-overexpressed transgenic Arabidopsis thaliana lines were acquired, followed by drought and high-temperature treatments, the plant phenotypes were observed, and the physiological and biochemical indexes were determined. Result RcFDH2 showed differential expression during the rehydration process of R. canescens; the coding sequence of RcFDH2 was successfully cloned, and multiple RcFDH2-overexpressed transgenic A. thaliana lines were obtained. It was found that under drought and high-temperature treatments, the RcFDH2 transgenic lines showed stronger tolerances to drought and high-temperature compared to wild-type plants. The physiological and biochemical indicators of RcFDH2-overexpressed transgenic A. thaliana and wild-type plants revealed that in the transgenic plants, the contents of three osmotic regulatory substances, the total chlorophyll content, and the activities of three antioxidant enzymes were significantly higher than those in the wild-type plants, while the malondialdehyde content was lower than that in the wild-type plants. Conclusion RcFDH2 can respond to desiccation and high-temperature stress treatments in R. canescens, and its overexpression probably enhanced the drought and high-temperature tolerance of A. thaliana maybe by regulating osmotic regulatory substances and antioxidant enzyme activities.

Objective To explore the chloroplast genome characteristics and phylogenetic relationships of Chrysanthemum cultivars of different cultivation types, providing a theoretical basis for research on the phylogeny, species identification and germplasm conservation of the genus Chrysanthemum. Method Chloroplast genome sequencing, assembly and annotation were performed on the groundcover Chrysanthemum × morifolium ‘Bei Lin Qiu Yun’, the single-headed cut flower C. morifolium ‘Sheng Xue’, and the traditional large-flowered C. morifolium ‘Fen Shi Ba’. Analysis of genomic structure, simple sequence repeats (SSRs), codon usage bias, inverted repeat (IR) boundary regions and collinearity characteristics was conducted. For phylogenetic analysis, nine chloroplast genome sequences, including eight from Chrysanthemum species and one outgroup from the Asteraceae family, were downloaded from GenBank. Result The chloroplast genomes of the three cultivars all exhibited a typical double-stranded circular quadripartite structure, ranging in length from 151 060 to 151 073 bp, with a GC content of 37.5%. A total of 131 genes were annotated in each genome, including 86 protein-coding genes, 37 transfer RNA (tRNA) genes, and 8 ribosomal RNA (rRNA) genes. Whole-genome alignment revealed high overall conservation of the chloroplast genomes: the IR regions showed higher conservation than the single-copy (SC) regions, and the non-coding regions had a higher mutation frequency than the coding regions, with no large-scale gene rearrangements detected. Boundary gene types were consistent across all samples. C. morifolium ‘Sheng Xue’ and C. morifolium ‘Fen Shi Ba’ each contained 68 SSRs, whereas C. morifolium ‘Bei Lin Qiu Yun’ contained 70 SSRs. Mononucleotide repeats (A/T-type) dominated the SSRs in all three cultivars, with no hexanucleotide repeats detected. Codon usage bias was weak (CAI: 0.168‒0.169; ENC: 49.127‒49.420), with a preference for A/U-ending codons. Selection pressure was identified as the key factor influencing codon usage bias. Phylogenetic analysis showed that C. morifolium ‘Sheng Xue’ and C. morifolium ‘Fen Shi Ba’ were closely related to cultivated C. morifolium, while C. morifolium ‘Bei Lin Qiu Yun’ was closely related to wild Chrysanthemum species. Conclusion The three Chrysanthemum cultivars of different cultivation types have a conserved chloroplast genomic structure and gene content. The groundcover C. morifolium is more closely related to wild species, whereas the traditional large-flowered C. morifolium and single-headed cut C. morifolium are more closely related to cultivated varieties.

Objective To elucidate the molecular regulatory mechanisms underlying the overwintering of ‘Zezhou’ celery in open fields, providing theoretical support for utilization of valuable germplasm resource and molecular breeding for cold tolerance in crops. Method Using ‘Duoliwang’ celery (CRY) at the seedling stage as the control, differentially accumulated metabolites and differentially expressed genes in the roots were analyzed between different varieties and different stages (seedling stage ZRY and flowering stage ZR) of ‘Zezhou’ celery. Result Metabolomic analysis revealed 57 significantly differentially accumulated metabolites in ZRY vs. CRY, and 187 in ZR vs. ZRY. Further screening identified 9 cold-tolerance-related differential metabolites, including phenylpropanoids and polyketides, lipids and lipid-like molecules, and organo-oxygen compounds, all of which showed increased accumulation. Transcriptomic analysis identified 2 988 differentially expressed genes in ZRY vs. CRY, with 1 001 upregulated and 1 987 downregulated. In ZR vs. ZRY, 8 869 differentially expressed genes were identified, comprising 4 153 upregulated and 4 716 downregulated genes. These genes were primarily enriched in pathways such as plant hormone signal transduction. Following overwintering in open fields, the content of the differential metabolite salicylic acid increased in ‘Zezhou’ celery. Analysis of the salicylic acid biosynthesis pathway indicated that the gene Ag2G02930 plays a significant role in its synthesis. Conclusion ‘Zezhou Celery’ resists cold stress by accumulating key metabolites such as salicylic acid and activating plant hormone signal transduction pathways. The salicylic acid synthesis-related gene Ag2G02930 plays a core regulatory role in its resistance to cold during overwintering in the open field.

Objective To address the severe phosphorus (P) immobilization and low plant availability in slightly alkaline soils, this study aimed to screen high-efficiency and stress-tolerant phosphate-solubilizing bacteria (PSB), optimize their culture conditions, clarify the P-solubilizing mechanism, and verify their effects on soil P activation and wheat growth promotion. Method A total of 26 candidate strains from 7 genera were used as test materials. High-efficiency PSB were isolated via primary screening by the plate transparent zone method (using inorganic P selective medium, with phosphorus-solubilizing index as the evaluation criterion) and secondary screening by the liquid culture method (soluble P content determined by molybdenum antimony anti-spectrophotometry). Single-factor experiments and response surface methodology (RSM) were conducted to optimize the cluture conditions. High-performance liquid chromatography (HPLC) was used to identify organic acid components, and Pearson correlation analysis was performed to reveal the relationship between organic acid profiles and P-solubilizing efficiency. Wheat pot experiments were carried out to evaluate soil application effects. Result An efficient PSB strain was identified as Enterobacter asburiae N4129-2AT. This strain maintained stable P-solubilizing capacity under low temperature (15 ℃) and high alkalinity (pH 10). After RSM optimization, the optimal culture conditions were confirmed to be 35 ℃, initial pH 6, and an inoculum size of 7.2%. Under these conditions, the soluble P content in fermentation broth reached 823.471 mg/L within 72 h, which was 23.7% higher than that before optimization, and the culture cycle was shortened by 96 h. E. asburiae N4129-2AT stably secreted six types of small-molecule organic acids. The core P-solubilizing mechanism was acidification mediated by organic acids, and the key factor improving P-solubilizing efficiency was organic acids strongly associated with P solubilization rather than their content proportion; total organic acid content only played an auxiliary role. In wheat pot experiments, all PSB treatments significantly increased soil available P content and wheat growth parameters compared with the control. The medium-dose optimized whole bacterial broth treatment showed the best performance: soil available P content increased by 376.76%, wheat the root dry weight increased by 157.4%, and root-shoot ratio decreased by 22.69%. A highly significant positive correlation was observed between soil available P and wheat growth parameters. Conclusion E.asburiae N4129-2AT possesses excellent stress tolerance (low temperature and high alkalinity) and high P-solubilizing efficiency. Its P-solubilizing mechanism, dominated by strongly associated organic acids through targeted optimization of organic acid profiles, breaks the traditional cognition that total organic acid content determines P-solubilizing capacity, providing a theoretical reference for the improvement of similar strains. This strain and its optimized culture conditions offer reliable technical support for P activation in slightly alkaline soils and high-yield wheat production.

Objective Strain GNLJ-7 was isolated from tomato rhizosphere. Its functional mechanisms were elucidated through whole-genome sequencing, aiming to provide an excellent microbial strain and theoretical support for developing green plant-growth-promoting and biocontrol agents. Method The effects of GNLJ-7 on tomato growth and Phelipanche aegyptiaca parasitism were evaluated via in-plate seed germination and pot experiments. Its phosphate-solubilizing, nitrogen-fixing, and siderophore-producing abilities were assessed using plate-based assays. Taxonomic identification of strain GNLJ-7 was performed by 16S rRNA and whole-genome sequencing. The genome was sequenced using both PacBio and Illumina platforms, followed by functional annotation via COG, KEGG, and GO databases. Secondary metabolite biosynthetic gene clusters and key metabolic pathways were predicted. Result Strain GNLJ‑7 was identified as Gordonia didemni. Pot experiments showed that the strain significantly promoted tomato root development, increasing root length and root weight by 26% and 47%, respectively, and effectively inhibited the germination and parasitism of P. aegyptiaca seeds. At a concentration of 1.0×10² CFU/mL, the inhibition rate on Orobanche seed germination reached 86.47%. Functional tests confirmed its abilities in nitrogen fixation, phosphate solubilization, and siderophore production. Key genes involved in indole‑3‑acetic acid (IAA) biosynthesis (trpA/B/C/D/E/F/S), phosphate solubilization (phoA, phoD), siderophore synthesis (pksD/E/F), and nitrogen fixation (nifH/A) were identified in the genome. Ten secondary metabolite biosynthetic gene clusters and metabolic pathways associated with plant‑growth‑promoting and biocontrol functions were predicted. Conclusion The strain Gordonia didemni GNLJ-7 significantly promotes tomato root growth and inhibits the parasitism of Phelipanche aegyptiaca. Genes associated with nitrogen fixation (nifH, nifA), phosphorus solubilization (phoA, phoD), siderophore synthesis (pksD/E/F), and IAA biosynthesis (trpA/B/C/D/E/F/S) identified in its genome collectively form the molecular basis for its plant growth-promoting and biocontrol activities.

Objective This study aims to elucidate the role of the soybean transcription factor GmWRKY23a in regulating root colonization by the beneficial bacterium Pseudomonas chlororaphis IRHB3. Methods Using the soybean cultivar Glycine max cv. Nandou 12, we systematically investigated the structural features of the GmWRKY23 gene and its encoded protein through a combination of bioinformatic analysis, molecular biology approaches, and the soybean hairy root-based transient genetic transformation. The regulatory mechanism of GmWRKY23a in response to IRHB3 root colonization was further characterized. Results Four gene sequences annotated as GmWRKY23 were retrieved from the NCBI and Phytozome databases. All encoded proteins contained the conserved WRKY domain and localized in the nucleus, but exhibited variations in protein structure and promoter cis-acting element composition. Among them, GmWRKY23a showed distinct tissue-specific expression and was significantly induced by IRHB3, the pathogen Fusarium oxysporum B3S1, phytohormones (SA, BR, MeJA), and salt stress (NaCl). Functional assays revealed that overexpression of GmWRKY23a promoted IRHB3 colonization in soybean roots. GmWRKY23a positively regulated the expression of key isoflavone biosynthetic genes, including chalcone synthase (CHS8), chalcone reductase (CHR1), and isoflavone synthase (IFS1), alleviated the transcriptional repression of IFS1 caused by IRHB3 colonization, and synergistically enhanced the expression of CHS8 and CHR1 in coordination with IRHB3. Conclusion GmWRKY23a positively modulates root colonization by the beneficial P. chlororaphis viatranscriptional activation of the isoflavone biosynthesis pathway. These findings provide new insights into the coordinated regulation of transcription and secondary metabolism in plant-beneficial microbe interactions.

Objective This study aimed to evaluate the antagonistic activity of Bacillus subtilis JY-7-2 against the Aconitum carmichaelii Debeaux root rot pathogen Fusarium oxysporum and its biocontrol efficacy against the root rot in the field, with genomic analysis to elucidate the underlying antimicrobial mechanisms. Method We evaluated the antagonistic activity of strain JY-7-2L with a plate confrontation assay. In field trials, we assessed its biocontrol efficacy against root rot and its effects on the growth of A. carmichaelii Debeaux. We performed whole-genome sequencing using the Illumina NovaSeq and Oxford Nanopore platforms, conducted functional annotation of the genome with the COG, CAZy, GO, and KEGG databases, and predicted secondary metabolite biosynthetic gene clusters with antiSMASH. Result Strain JY-7-2L significantly inhibited the growth of F. oxysporum and reduced the incidence of root rot in the field, achieving a biocontrol efficacy of 89.26%-93.77%. It also increased the dry weights of plant stems, main roots, and lateral roots. Both the cell-free fermentation filtrate and volatile organic compounds of JY-7-2L markedly suppressed the growth of F. oxysporum. The genome of JY-7-2L was 4.19 Mb in length with a GC content of 43.57%, encoding 4 234 genes. Genomic analysis revealed the presence of genes associated with antibiotic synthesis, biofilm formation, pathogen inhibition, plant growth promotion, and induction of plant systemic resistance. AntiSMASH analysis identified 11 secondary metabolite biosynthesis gene clusters, including 6 involved in the synthesis of antimicrobial compounds, and 3 potentially contribute to novel bioactive metabolites. Conclusion With its dual biocontrol and plant growth-promoting activities, Bacillus subtilis JY-7-2L shows strong potential for development as a biofertilizer and biocontrol agent.

Objective This study aimed to isolate and screen salt-alkali tolerant plant growth-promoting rhizobacteria, thereby providing microbial resources and theoretical foundations for the development of functional microbial fertilizers and the remediation of saline-alkali soils. Method Rhizosphere soil samples were collected from cotton plants in Yingjisha County, Xinjiang. Salt-alkali tolerant bacteria were isolated and screened using the serial dilution method. The strains were identified by 16S rRNA gene sequencing, and their taxonomic status was determined based on morphological, physiological, and biochemical characteristics. Salt-alkali tolerance of the strain was assessed using media with NaCl concentrations of 0–200 g/L and pH values of 4.0–13.0. Plant growth-promoting traits, such as phosphate solubilization, siderophore production, nitrogen fixation, biofilm formation, and amylase and cellulase activities, were evaluated using specific functional media. Finally, the effect of the strain on tomato seedling growth was examined through plate and pot experiments. Result We isolated a salt-alkali tolerant strain. 16S rRNA gene sequence analysis showed that the strain shared 99.72% similarity with the type strain Bacillus cabrialesii TE3^T, and we identified it as B. cabrialesii zp-4245. Strain zp-4245 could grow under conditions of 80 g/L NaCl and pH 9.0, and exhibited multiple plant growth-promoting traits, including siderophore production, nitrogen fixation, and the production of amylase, cellulase, and biofilm. In plate assays, we found that strain zp-4245 significantly promoted the growth of tomato seedlings under different salt-alkali stress conditions. Compared with the control at pH 5.8, inoculation with the strain increased lateral root number, aboveground fresh weight, underground fresh weight, and total fresh weight by 284.3%, 87.3%, 118.4%, and 94.2%, respectively. At pH 8.0, the increases were 361.7%, 92.3%, 141.7%, and 101.9%, respectively. Under combined stress of pH 8.0 and 2.925 g/L NaCl, the lateral root number increased by 707.6% compared to the control. Pot experiments demonstrated the growth-promoting effects of strain zp-4245 on tomato plants. Under non-salt-stress conditions, inoculation with strain zp-4245 increased aboveground fresh weight and plant height by 11.7% and 13.9%, respectively. Under salt stress (6 g/kg NaCl), inoculation increased underground fresh weight and plant height by 28.6% and 10.7%, respectively, compared with the control. Conclusion B. cabrialesii zp-4245 exhibits excellent salt-alkali tolerance and plant growth-promoting capabilities, significantly enhancing tomato seedling growth under varied saline-alkaline conditions. These findings provide valuable microbial resources and a theoretical foundation for developing microbial fertilizers for saline-alkali soils improvement.

Objective The evolved strain Alk_55 was obtained from Rhizobium yanglingense CCBAU 01603 through continuous subculturing. This work aims to investigate the underlying reasons for the significant enhancement of salt-alkali tolerance caused by SNP changes in aotM and trmD in the Alk_55 genome. Method The regulation of salt-alkali tolerance, cell membrane damage, intracellular antioxidant enzyme activity, and reactive oxygen species (ROS) content in rhizobia were assessed, based on two mutants of rhizobia, △aotM and △trmD, created by deleting genes aotM and trmD respectively. Result Mutant △aotM and △trmD had stronger salt-alkali tolerance and reduced sensitivity to salt-alkali stress within a certain concentration range, compared with the wild-type strain. Significantly lower malondialdehyde (MDA) content indicated less lipid peroxidative damage to the cell membrane. Intracellular superoxide dismutase (SOD) and catalase (CAT) activities markedly increased, whereas ROS content significantly decreased. Conclusion Genes aotM and trmD strongly regulate salt-alkali tolerance, intracellular antioxidase activity, and oxidative stress-induced cell membrane damage in rhizobia.

Objective The supply form of inorganic carbon (Ci) is a critical environmental factor regulating photosynthetic efficiency in cyanobacteria. However, the inorganic carbon utilization preference of cyanobacteria at the single-cell level and the corresponding molecular regulatory mechanism remain unclear under the coexistence of CO₂ and \mathrm{H}\mathrm{C}{\mathrm{O}}_{\mathrm{3}}^{-}. This study used the model strain Synechococcus sp. PCC7942 to reveal its utilization preference for different inorganic carbon sources at the single-cell level. Method This study integrated ¹³C stable isotope probing, single-cell Raman spectroscopy (Raman-SIP), and transcriptome sequencing. Six carbon source treatments were set, including ambient ¹²C-NaHCO₃, ¹³C-NaHCO₃, air, ¹²C-CO₂, ¹²C-CO₂ + ¹³C-NaHCO₃, and ¹³C-CO₂ + ¹²C-NaHCO₃. Cells were cultured in BG11 medium under controlled light and temperature. The ¹³C assimilation and carbon source utilization characteristics were quantitatively characterized by detecting the shift of the carotenoid v1 peak using single-cell Raman spectroscopy. Transcriptome sequencing was performed on samples grown on single carbon sources (¹²C-NaHCO₃ and ¹²C-CO₂). The regulatory mechanism of carbon metabolism pathways was dissected through data quality control, genome alignment, screening of differentially expressed genes, and GO/KEGG enrichment analysis. Result In the CO₂/\mathrm{H}\mathrm{C}{\mathrm{O}}_{\mathrm{3}}^{-} coexistence system, the strain predominantly utilizes CO₂—an observation quantitatively confirmed by dynamic redshifts in the Raman carotenoid v1 peak. RNA-seq analysis revealed 2 750 differentially expressed genes (DEGs) with annotations in the CO₂-treated group, among which 1 083 were significantly differentially expressed. Most genes encoding enzymes of the Calvin-Benson cycle showed downregulated transcription, whereas rbcLS expression was upregulated. Conclusion This study integrates single-cell Raman spectroscopy and transcriptomics to demonstrate that Synechococcus sp. PCC7942 preferentially utilizes CO₂ and clarify its transcriptional regulatory mechanism. This work establishes a systematic linkage among carbon source preference, Raman spectral characterization, and transcriptional regulation at the single-cell level.