Stem-reserved water-soluble carbohydrates (WSC) serve as a crucial carbon source for grain filling in wheat (Triticum aestivum). Enhancing stem WSC content is a key strategy for achieving high and stable grain yield. The major-effect QTL QWSC.caas-7DS, which regulates stem WSC content at 10 d after flowering (WSC10), was previously identified in the 'Yangmai 16' (YM16)/'Zhongmai 895' (ZM895) doubled haploid (DH) population, with physical interval of 11.7 Mb. To fine-map this QTL, identify the candidate genes, and develop molecular markers, a cross XY015/XY102 was made with 2 DH lines XY015 and XY102 that contrast in genotype at QWSC.caas-7DS but shared identical genotypes at other known WSC loci. Subsequently, F₂, F_2:3, and F_2:4 segregating populations were constructed. Based on whole-genome resequencing data of YM16 and ZM895, 20 kompetitive allele-specific PCR (KASP) markers were successfully developed within the target interval, and a high-density genetic map was constructed using 755 F₂ individuals. The homozygous F_2:4 lines derived from 35 recombinant F₂ plants were used for phenotyping of WSC10 in 3 environments; in combination with the genotypic data of these lines, QWSC.caas-7DS was finely mapped to the 66.44~69.14 Mb interval on chromosome 7DS. Comprehensive analysis of genomic resequencing data of YM16 and ZM895, along with annotations and expression profiles of the genes in the QTL interval, 6 genes were preliminarily presumed as the candidate genes for QWSC.caas-7DS. According to the gene expression profile database, Vrn-D3 (gene ID: TraesCS7D01G111600), one of the candidate genes, showed high expression levels in stem and leaf tissues, and it may regulate stem WSC and thousand-kernel weight (TKW) by modulating carbon balance between source and sink tissues. Vrn-D3 was cloned from YM16 and ZM895, and a G base "insertion/deletion" variation located in the third exon was identified. A molecular marker, Vrn-D3M, was developed for genotyping this variation and was validated in 157 wheat varieties from the Yellow and Huai Valleys Winter Wheat Zone. The results indicated that Vrn-D3M could be effectively used for molecular breeding targeting for improvement of WSC10 and TKW. The findings of this study lay a solid foundation for the final cloning and functional study of QWSC.caas-7DS, and also provide an efficient molecular tool for breeding applications.

Grain filling rate (GFR) is a key factor influencing wheat (Triticum aestivum) thousand-kernel weight (TKW) and yield, while cell wall invertases (CWINVs) are core enzymes regulating plant carbohydrate metabolism and grain filling. It is of high significance to explore variations of wheat CWINV family genes and develop molecular markers for genetic improvement of wheat GFR and yield. In this study, a natural population comprising 166 wheat varieties (advanced lines) from the Yellow and Huai Valleys Winter Wheat Zone was used to measure the maximum grain filling rate (GFR_max) and TKW under 4 environments. With the genotyping data of wheat 660K and 90K SNP chips, genome-wide association study (GWAS) was conducted and a major-effect QTL simultaneously associated with GFR_max and TKW was identified at the 60.60~62.33 Mb interval on chromosome 4B, explaining 7.48%~12.40% and 7.53%~9.50% of the phenotypic variance for GFR_max and TKW, respectively. Referencing the 'Chinese Spring' reference genome, the TaCWINV41 gene (TraesCS4B02G066000) belonging to the CWINV family was located within the interval of this QTL. TaCWINV41 gene was cloned from cultivars 'Yangmai 16' and 'Zhongmai 895', and 2 SNPs were identified in its third exon; one of which, at position 1 892, was a C/T missense mutation. Based on this SNP, a kompetitive allele-specific PCR (KASP) marker, TaCWINV41_KASP, was developed. The reliability of this marker was validated using GFR_max and TKW phenotypic data of 194 wheat varieties. t-test analysis revealed that varieties with TT genotype exhibited significantly higher GFR_max and TKW than those with CC genotype (P<0.05, P<0.01, or P<0.001), confirming the value of TaCWINV41_KASP for molecular breeding. In summary, this study identified a major QTL simultaneously regulating wheat GFR and TKW through GWAS, uncovered allelic variation in the candidate gene TaCWINV41 at this locus, and developed a KASP marker for molecular breeding of wheat GFR and TKW. These findings provide theoretical insights into the genetic regulation of wheat GFR and offer an efficient molecular tool for genetic improvement of GFR and grain yield.

In common wheat (Triticum aestivum), the black awn trait is relatively rare, and its genetic mechanism remains unclear. This study aims to genetically map the black awn trait in wheat, laying the foundation for subsequent gene cloning and breeding applications. Analysis of the F_2∶9 RIL population derived from 'Yang16G216' (white awn)/ 'Yang16M6393' (black awn) revealed a segregation ratio of white to black awns consistent with 3∶1. Further bulked segregant exome sequencing (BSE-Seq) was performed on the F_2∶5 RIL population from the cross 'Yang16C106' (white awn)/'Yang20M5623' (black awn). Among 86 566 single nucleotide polymorphism (SNP) sites, 456 ones were highly significantly associated with the black awn trait (at the 99% confidence level). Of these, 99.34% (453) were densely distributed in the distal end of chromosome 1AS (65) and the peri-centromeric region of chromosome 1BL (388), demonstrating that the trait was co-regulated by these 2 loci. Fine mapping localized the chromosome 1AS locus to the 0~8.1 Mb interval, and the chromosome 1BL locus to the 360.4~399.5 Mb interval, which was found to be closely linked with the stripe rust resistance gene Yr26. By breaking this linkage and combining multi-disease resistance gene detection, 3 white-awn lines were selected from the F_2∶5 RIL population, which pyramided the stripe rust resistance gene Yr26, powdery mildew resistance gene Pm21, leaf rust resistance gene LrYang16G216, and Fusarium head blight resistance gene Fhb1. This study provides both theoretical and material foundations for map-based cloning of the black awn gene, as well as for pyramiding multiple disease resistances and molecular breeding applications.

Wheat flour pasting properties are important indicators of processing quality. To dissect the genetic basis of these traits, in this study, 221 wheat (Triticum aestivum) accessions were evaluated for 7 wheat flour pasting traits, including peak viscosity, trough viscosity, breakdown, final viscosity, setback, peak time, and pasting temperature, across 4 environments over 3 years. Using 90K SNP array data, a genome-wide association study (GWAS) was conducted with 5 models, namely the general linear model (GLM), mixed linear model (MLM), multi-locus mixed linear model (MLMM), fixed and random model circulating probability unification (FarmCPU), and bayesian-information and linkage-disequilibrium iteratively nested keyway (BLINK). All 7 traits showed abundant phenotypic variation, with coefficients of variation ranging from 0.92% to 33.02% and broad-sense heritability ranging from 40.31% to 70.36%. Joint GWAS analysis across multiple models identified 8 significant and stable association loci on chromosomes 2B, 5A, 6B, and 7A, each explaining 2.09% to 9.26% of the phenotypic variation. Haplotype analysis further identified the superior haplotypes of these high-confidence loci and their distribution frequencies in the population. Candidate genes were screened within the 200 kb regions upstream and downstream of the high-confidence loci. Combined with gene expression analysis, 2 candidate genes related to wheat flour pasting properties were identified, namely TraesCS6B03G0474900 (encoding a glycine cleavage system H family protein) and TraesCS2B03G0671500 (encoding a 40S ribosomal protein S12). This study provides a reference for the genetic dissection and molecular breeding of wheat flour quality.

Starch branching enzyme (SBE) is a type of glycosyltransferases that has 2 catalytic functions, it synthesizes and converts amylopectin as part of the starch synthesis pathway. It plays a key regulatory role during grain development in wheat (Triticum aestivum). To investigate the expression characteristics of TaSBE family genes during wheat grain development and identify superior haplotypes, bioinformatics methods were used to identify TaSBE family members across the genome, analyze their expression patterns, and develop and validate functional molecular markers. The results showed that 15 TaSBE genes were found throughout the wheat genome, dispersed on chromosomes 2A, 2B, 2D, 7A, 7B, and 7D. These genes could be divided into 3 subfamilies, with members of the same subfamily sharing comparable gene structures and shared motifs. Intraspecific synteny analysis revealed 9 pairs of chromosomes with intragroup synteny, demonstrating the evolutionary conservation of family members. Analysis of cis-acting elements revealed that TaSBE family genes contain a large number of developmental response elements. RNA-seq and qPCR analysis revealed that the TaSBE1-7A, TaSBE1-7B, and TaSBE1-7D genes were highly expressed in grains. Sequence polymorphism analysis revealed the presence of regular SNP sites in the sequences of TaSBE1-7A and TaSBE1-7B, allowing them to be classified into 2 haplotypes. KASP molecular markers were created for TaSBE1-7A and TaSBE1-7B. The results demonstrated that the KASP marker, positioned at 1 088 bp (G/A) in the TaSBE1-7B complete genome sequence, efficiently discriminated the 2 haplotypes. Associative analysis of grain starch content revealed that varieties with the TaSBE1-7B-HapⅠ haplotype had considerably higher grain starch content than those with TaSBE1-7B-HapⅡ, and TaSBE1-7B-HapⅠ was under positive selection during wheat breeding. This study provides a theoretical foundation for further investigation into the molecular processes by which the TaSBE1 gene controls grain growth and starch synthesis.

Deep analysis of carotenoid contents and key gene expression patterns in wheat (Triticum aestivum) grains is of great significance for breeding wheat varieties with high carotenoid content. In this study, carotenoid components in grains of 3 yellow-flour varieties (JM1, JM1803, and JM8040) and 3 white-flour varieties (JN17, JM22, and JM44) were comparatively analyzed using high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS/MS). Results showed a total of 7 xanthophylls, 10 xanthophyll esters and 3 carotenes were identified across the 6 varieties. 8 components, including lutein and zeaxanthin, were widely detected in all varieties. JM1, JM1803, and JM8040 contained more components, including 4 unique components such as violaxanthin, lutein dimyristate, β-carotene, and phytoene. Quantitative analysis revealed that total carotenoid content in JM1, JM1803, and JM8040 reached 9.30, 8.97, and 5.67 μg/g, respectively, significantly higher than other varieties (P<0.05). These 3 varieties also exhibited significantly higher total xanthophylls, total xanthophyll esters, total carotenoids, and most of carotenoid components. Further analysis of the expression patterns of key carotenoid biosynthesis genes (PSY1, PDS, ZDS, LCYE, and LCYB) in developing grains showed that although the expression trends of the 5 genes were consistent across varieties, expression levels of PSY1, PDS, and ZDS in JM1, JM1803, and JM8040 were significantly higher at 14 and 21 d after flowering (DAF)(P<0.05). Additionally, expression leve of LCYE in these varieties was significantly higher at 7 DAF, and expression level of LCYB was significantly elevated at 7, 14, and 28 DAF (P<0.05). Moreover, these gene expression levels exhibited an extremely significant positive correlation with contents of total carotenoid, xanthophylls, xanthophyll esters, and carotenes (P<0.01). The high expression of these key genes may contribute to the elevated carotenoid content in these 3 yellow-flour varieties. This study provides a theoretical foundation for understanding carotenoid biosynthesis pathways and breeding wheat varieties with enhanced carotenoid content.

In conventional hybrid breeding of wheat (Triticum aestivum), parental materials primarily consist of currently promoted varieties. This tendency toward similar parental sources has led to an increasingly narrow genetic foundation in newly developed cultivars and heightened genetic homogeneity among varieties. Consequently, valuable alleles for disease resistance, cold tolerance, and broad-spectrum stress tolerance—abundantly present in local germplasm resources—are being lost. A thorough analysis of the genetic diversity and phylogenetic relationships between Ningxia spring wheat landraces and cultivated varieties holds significant implications for optimizing parent selection strategies in breeding programs and enhancing progeny selection efficiency. This study employed 100K-GBTS chips for genotyping analysis of 314 spring wheat germplasm resources. Cluster analysis was conducted based on genetic distance, and the 314 wheat accessions were classified into 3 clusters. Further functional gene analysis revealed that '96NS4019' and '14Y1269' exhibited the best overall performance in terms of grain weight and grain size; three accessions—'M423', '91 Yun 315', and 'MJ317'—expressed all 5 dwarf genes. Analysis of gene combinations and single-factor variance of related traits revealed that materials with the Rht8+Rht-D1 genotype exhibited significantly lower plant height than those with the QPht-2D+Rht8+Rht-D1 genotype (P<0.05); Materials containing the TaCwi-A1+TaGS5-A1+TaT6P gene combination exhibited significantly higher thousand-kernel weight than those lacking high-grain-weight genes (P<0.05). These findings provide germplasm resources and molecular evidence for scientifically selecting parental combinations and directing genetic improvement of target traits in future spring wheat hybrid breeding programs in Ningxia.

WRKY24 plays an important role in regulating plant responses to abiotic stresses, particularly cold stress. However, limited information is available regarding the expression patterns of WRKY24 family members in wheat (Triticum aestivum) under low-temperature and 2,4-epibrassinolide (2,4-EBR) treatments. In this study, 6 members of the wheat WRKY24 family (TaWRKY24-1/2/3/4/5/6) were selected for systematic analysis using bioinformatic approaches and qRT-PCR. Their expression patterns were examined in strong tolerance to late spring cold stress wheat cultivar 'Ji Mai 22' and weak tolerance to late spring cold stress cultivar 'Ma Zha Mai'. The results showed that TaWRKY24-1/2/3 were located on chromosome 1 (1A, 1B, and 1D), whereas TaWRKY24-4/5/6 were located on chromosome 3 (3A, 3B, and 3D), and these 2 groups exhibited significant differences in predicted molecular weight and isoelectric point of the encoded proteins. Phylogenetic analysis revealed that wheat WRKY24 proteins were most closely related to rice (Oryza sativa) homologs, and that members within the same subfamily displayed distinct differences in motif composition and gene structure. Promoter analysis identified abundant cis-acting regulatory elements, including those responsive to abscisic acid (ABA), gibberellin (GA), methyl jasmonate (MeJA), light, low temperature, and drought stress, indicating that TaWRKY24 genes were regulated by multiple hormonal and environmental cues. qRT-PCR analysis showed that under low-temperature treatment, transcript levels of TaWRKY24-1 increased in both 'Ji Mai 22' and 'Ma Zha Mai', whereas TaWRKY24-2/3/4/6 exhibited reduced expression in both cultivars. Under combined low-temperature and 2,4-EBR treatment, transcript levels of TaWRKY24-1/3/4/5/6 were further elevated in both cultivars. These results revealed that the TaWRKY24 genes were regulated by various hormones and environmental factors, and under low-temperature stress and 2,4-EBR treatment, the expression patterns of different TaWRKY24 genes varied in strong and weak cold-resistant wheat varieties. This study provides a foundation for further elucidation of the molecular mechanisms underlying cold stress responses in wheat and for the potential utilization of WRKY24 family genes in cold-tolerance breeding.

The Triticum aestivum xylanase inhibitors (TaXIs) can suppress the enzymatic activity of the GH11 family xylanase from Fusarium graminearum and its induced cell necrosis, but the transcriptional regulatory mechanism of XIs remains unclear. In this study, the promoter sequence of the TaXI-Ⅲ gene was cloned from wheat cultivar 'Annong 1589', and its cis-acting elements were analyzed by PlantCARE database. The TaXI-Ⅲ promoter fragment was used as bait to screen the interacting transcription factors by yeast one-hybrid (Y1H) assay, and the binding specificity between transcription factors and promoter was confirmed by point-to-point Y1H, dual-luciferase reporter system, and GUS histochemical staining. The results showed that the promoter sequence of 2 138 bp upstream of TaXI-Ⅲ gene was cloned successfully, which contained cis-acting elements such as jasmonic acid response, biological/abiotic stress response, MYB and MYC binding sites. Four transcription factors were screened from wheat cDNA library with pHis-TaXI-Ⅲpro as bait vector and 120 mmol/L 3-amino-1,2,4-triazole (3-AT ) as selective agent, they were TaMYB30, TaPIEP1, TaTGA1a and TaWRKY33. Among them, TaWRKY33 was verified to bind to the W-box within the TaXI-Ⅲ promoter and regulate its expression. qPCR analysis revealed a co-expression pattern for TaWRKY33 and TaXI-Ⅲ under methyl jasmonate and F. graminearum treatment. Furthermore, overexpression of TaWRKY33 in Arabidopsis thaliana enhanced plant resistance to F. graminearum. These results indicated that TaWRKY33 might enhance resistance to F. graminearum by binding to the W-box in the TaXI-Ⅲ promoter and activating gene expression through the jasmonic acid signaling pathway. This study provides important reference information for further elucidating the transcriptional regulatory mechanism of the TaXI-Ⅲ gene.

The vesicle transport protein (SFT2) gene family plays a key role in membrane trafficking and stress responses in eukaryotes. However, its members in wheat (Triticum aestivum) and their functions under cadmium (Cd) stress remain unclear. This study performed a genome-wide identification of the SFT2 family in wheat, with focus on the biological function of TaSFT2 in Cd stress response. Results showed that 21 TaSFT2 members were identified in the wheat genome, distributed across 15 chromosomes and relatively evenly among the A, B, and D subgenomes. Phylogenetic analysis classified SFT2 proteins from wheat, maize (Zea mays), and Arabidopsis thaliana into 4 groups, with wheat TaSFT2s mainly clustered in groups D, indicating functional divergence during evolution. Gene structure and conserved motif analyses revealed similar structural characteristics among homologous genes, with all members containing 4~5 introns. Cis-acting element analysis identified multiple hormone-responsive elements (methyl jasmonate, abscisic acid) and abiotic stress-responsive elements (drought, low temperature, anoxia) in TaSFT2 promoters, suggesting their broad involvement in hormone signaling and stress responses. Expression profiling demonstrated tissue and development stage-specific expression patterns of TaSFT2s, with some members showing significant expression in spikes and roots, implying potential roles in spike development and root function, respectively. Heterologous expression assays confirmed that TaSFT2-1 overexpression significantly enhanced Cd stress tolerance in Escherichia coli. This study elucidated the member composition, evolutionary characteristics, and expression patterns of the wheat SFT2 gene family, as well as the biological function of TaSFT2-1 in response to Cd stress. These findings provide a theoretical basis and genetic resources for dissecting the mechanisms underlying heavy metal tolerance in wheat and for developing Cd-tolerant germplasm.

Improvement of disease resistance, quality, and high yield simultaneously has been a long-standing goal in wheat (Triticum aestivum) breeding. Breeding new high-yielding dryland wheat varieties with both stripe rust resistance and high quality is particularly important for developing modern cold-arid agriculture in Gansu province. In this study, 'Ta04-44-1-3' wheat line developed from recurrent selection population selection of Taigu genic male-sterile wheat carrying the known high-quality glutenin subunit genes Ax1/Ax2* and Bx7+By8, as well as the pleiotropic stripe rust resistance genes Lr34/Yr18/Pm38/Sr57 and Lr27/Yr30/Sr2/Sb3 was utilized as the female parent, and 'Lantian 19', which possessed the known high-quality glutenin subunits Bx7+By8, Bx7+By9, Glu-A3d, as the male parent for hybridization. Using specific molecular markers for the high-quality glutenin subunit and stripe rust resistance genes, combined with disease resistance evaluation, quality assessment, and field-based comprehensive trait selection, a new winter wheat variety named 'Lantian 214' was developed. This variety combines high-quality, stripe rust resistance, stress tolerance, and high yield, making it suitable for dryland conditions. The variety possessed the high-quality glutenin subunits Ax1/Ax2*, Bx7+By8, Bx7+By9, Glu-A3d, as well as the pleiotropic resistance genes Lr34/Yr18/Pm38/Sr57 and Lr27/Yr30/Sr2/Sb3. Disease resistance evaluation revealed that the variety was moderately susceptible (MS) to mixed races of stripe rust at the seedling stage but exhibited immunity to prevalent mixed races at the adult plant stage. Grain quality analysis indicated that its grain test weight was 829 g/L, protein content was 13.4%, wet gluten content was 33.2%, dough stability time was 6.8 min, water absorption rate was 58.8%, maximum resistance to extension was 426 E.U, and extensibility area was 92 cm², meeting the standards for medium-strong gluten wheat. The development of new winter wheat cultivar 'Lantian 214' provides a high-quality, disease-resistant winter wheat variety suitable for dryland production in the mountainous southeastern Gansu. It also offers new parental germplasm for future efforts in pyramiding multiple genes for breeding in this region.

Wheat (Triticum aestivum) leaf rust, caused by the airborne fungal pathogen Puccinia triticina, is a major threat to wheat production and global food security. Effective management of this disease requires the exploration of novel resistance sources, the cloning of resistance genes, the development of linked molecular markers, and the breeding of resistant cultivars through molecular breeding approaches. This review systematically summarizes recent advances in research on wheat resistance to leaf rust in China and worldwide. It covers the resistance levels of predominant Chinese wheat cultivars and currently nationally-approved varieties, the identification and cloning of resistance genes from common wheat and its wild relatives, as well as the application and distribution of major resistance genes in China. In addition, the review consolidates resistance gene resources that are effective against prevalent physiological races in the country. By synthesizing research progress in resistance identification, genetic analysis, gene mapping, and germplasm innovation, this work aims to provide a theoretical reference for breeding wheat varieties with broad-spectrum and durable resistance to leaf rust.

Oleosin (OLE) plays a crucial role in the synthesis and storage of plant oils. Using Paeonia delavayi as the experimental material, this study conducted gene cloning, bioinformatics analysis, expression pattern analysis, and subcellular localization analysis to initially clarify the possible role of PdOLE gene in the oil synthesis of P. delavayi. The results showed that the PdOLE gene (GenBank No. PX564745) sequence was 411 bp in length and could encode 136 amino acids. Bioinformatics analysis indicated that the protein contained an OLE superfamily domain and 13 phosphorylation sites, lacked a signal peptide, and exhibited the highest proportion of α-helix in its secondary and tertiary structures. Sequence alignment results showed that the PdOLE protein had relatively low identity with OLE homologs from the closely related species P. ostii and P. lactiflora. Notably, it showed significantly higher homology with OLE proteins derived from 4 representative oil crops: Theobroma cacao, Olea europaea, Sesamum indicum, and Helianthus annuus. Consistent with the results, phylogenetic tree construction further demonstrated that the PdOLE protein formed distinct clade separated from the OLE proteins of the closely related species mentioned above, and clustered more closely with those from the selected oil crops, indicating a closer evolutionary affinity. The qRT-PCR results indicated that the expression level of the PdOLE gene in seeds was significantly higher than that in other organs (P<0.05), reaching its peak at 90 DPF (day post flowers). Nile red staining of paraffin sections demonstrated that the number of seed lipid droplet increased significantly at mid-development (after 60 DPF), accounting well for the PdOLE expression trend. Subcellular localization results indicated that the PdOLE protein was localized to the cell membrane. This study would provide a scientific basis for future investigations aimed at elucidating OLE gene function and the oil biosynthesis pathway of P. delavayi.

Milk fat synthesis is a critical determinant of the nutritional and economic value of sheep milk, whose post-transcriptional regulatory mechanisms remain incompletely elucidated. This study was conducted to decipher the regulatory role of the competing endogenous RNA (ceRNA) network in milk fat synthesis within the mammary gland of sheep (Ovis aries). Mammary tissue samples were collected from Hu sheep during late pregnancy (14 d prepartum) and peak lactation (14 d postpartum) for whole-transcriptome sequencing. Bioinformatic analyses were performed to construct the ceRNA network and identify target relationships pertinent to lipid synthesis. A total of 1 471 differentially expressed mRNAs were identified between the 2 stages, with 647 upregulated and 824 downregulated during lactation. Among them, suppressor of cytokine signaling 3 (SOCS3), lipin 1 (LPIN1), and stearoyl-CoA desaturase 5 (SCD5) are closely associated with milk fat synthesis. A ceRNA network encompassing 3 mRNAs, 6 miRNAs, and 6 lncRNAs was successfully constructed. Specifically, 2 lncRNAs, MSTRG.15867.1 and XR_006057437.1, were significantly upregulated (P<0.01) in breast tissue at 14 d postpartum, coinciding with the upregulation of their target gene, SOCS3 (P<0.05), forming a putative regulatory axis. Another lncRNA, MSTRG.9360.1, was significantly upregulated (P<0.05) in breast tissue at 14 d postpartum and found to target LPIN1, forming 2 regulatory axes. Furthermore, the MSTRG.15301.43-miR-6715-SCD5 axis was predicted to operate via the ceRNA mechanism. Functional enrichment analysis revealed that the differentially expressed genes were predominantly involved in cell cycle and lipid metabolic pathways, suggesting a coordinated shift from cellular proliferation to active lipid synthesis during lactogenesis. These findings elucidate the involvement of ceRNA networks in regulating milk fat synthesis across different lactation stages in sheep, providing novel insights and potential molecular targets for the breeding of high lactation sheep.

Pseudocohnilembus persalinus is a pathogenic scuticociliate parasite in mariculture fish species. In this study, the alternative oxidase (AOX) gene from Pseudocohnilembus persalinus was cloned. The AOX gene family was systematically identified at the genomic level, its biological functions and expression characteristics were predicted and analyzed. The full-length cDNA of the alternative oxidase 3 (AOX3) gene from P. persalinus was cloned using the rapid amplification of cDNA ends (RACE) technique. The AOXs gene family members in P. persalinus was systematically identified by screening the genome using the conserved Pfam domain PF01786. Bioinformatics approaches were applied to analyze the physicochemical properties, subcellular localization, gene structure, Scaffold localization, promoter cis-regulatory elements, and phylogenetic relationships of these genes. Expression analysis of P. persalinus AOXs gene family members were carried out by qRT-PCR during the trophic and cyst stages and under temperature stress. The result showed that the full-length cDNA of the AOX3 gene from P. persalinus was cloned, with a total length of 1 091 bp (GenBank No. PV953026), including a 47 bp 5'-UTR, a 150 bp 3'-UTR, and an ORF of 894 bp. Four AOXs gene family members (designated as PpAOX1~PpAOX4) were identified in the whole genome; All PpAOXs were predicted to be hydrophilic proteins without signal peptides and were primarily localized in the cytoplasm and mitochondria. Cis-acting regulatory element analysis of the promoter regions suggested that PpAOXs should be involved in responses to hypoxia, temperature stress, and defense-related stimuli. Phylogenetic analysis of 33 AOXs genes from 13 protist species revealed that the 4 AOXs members of P. persalinus clustered within a clade of Oligohymenophorean ciliates. qRT-PCR results showed that PpAOX1 was highly expressed during the trophic stage, whereas PpAOX2~PpAOX4 exhibited significant upregulation during the cyst stage. The PpAOXs gene family members displayed distinct expression patterns in response to temperature stress. This study provides critical insights for further investigating the biological functions of PpAOXs and deciphering the adaptive strategies of P. persalinus in response to environmental stress.

Potato (Solanum tuberosum) common scab, caused by pathogenic Streptomyces spp., is an important soil-borne disease that occurs worldwide. Non-pathogenic Streptomyces can produce a large number of metabolites with antimicrobial and growth-promoting effects, making them one of the microorganisms with the greatest biocontrol potential. To obtain non-pathogenic Streptomyces strains with biocontrol potential against Streptomyces scabies which causes potato common scab, this study conducted series of assays, including screening for antimicrobial activity of single strain in vitro, preparation for antagonistic Streptomyces combinations, testing for antimicrobial activity of the combinations, molecular identification of strains included in the different combinations, analysis for their antimicrobial and growth-promoting characteristics, and evaluation for biocontrol efficacy in greenhouse. The results showed that 15 strains with significant antagonistic effects against the S. scabies strain LBX-7 were obtained from 77 non-pathogenic Streptomyces strains. Among them, strain No. ZL21 exhibited the widest inhibition band (8.17 mm). After strain compatibility testing and combinations preparation, 3 combinations with antagonistic effects against LBX-7 were obtained. Among these, combination No. C (ZL21\ZL109\ZL144) showed the best antimicrobial effect, with an inhibition band of 7.67 mm in width. Multi-gene sequencing were used to identify the strains in the 3 combinations, strains No. ZL21 and ZL144 belonged to S. olivaceus, while strains ZL54, ZL68, ZL79, ZL90, ZL109 and 05B3 were identified as S. parvus. All strains in the combinations had the abilities to produce protease and chitinase, decompose potassium and fix nitrogen, but none of them could dissolve inorganic phosphorus. Several strains were capable of producing indole-3-acetic acid, cellulase, laccase and amylase. Greenhouse pot control experiment indicated that ZL21 exhibited the highest biocontrol efficacy (75.2%) in all the tested single strains, and combination No. C showed the best efficacy (54.4%) among the 3 combinations in greenhouse. This study obtained candidate Streptomyces strains ZL21 with good biocontrol efficacy against potato common scab. This strain exhibits promising potential as resources for controlling potato common scab.

Ultra-low temperature cryopreservation is essential for establishing a poultry germplasm bank. The establishment of poultry germplasm cryobanks faces multiple challenges, including the inherent cryosensitivity of spermatozoa, the inability to successfully cryopreserve oocytes, reduced post-thaw viability, immature genetic material resuscitation technologies, and limited survival of offspring. Currently, ultra-low temperature cryopreservation technologies for poultry genetic resources remain underdeveloped, with core issues such as ultra-low temperature cryopreservation efficacy and underlying freezing mechanisms requiring further in-depth investigation. In this paper, advances in the cryopreservation of semen, somatic cells, primordial germ cells (PGCs) and gonadal tissues of poultry were reviewed, and the findings were summarized to provide a reference for the conservation of poultry germplasm resources.