Domestic yaks are a crucial livestock species on the Qinghai-Tibet Plateau and its surrounding regions, providing indispensable resources for local residents' livelihoods and production. China harbors the world's largest yak population and possesses exceptionally rich yak genetic resources. In this study, we performed whole-genome resequencing on 200 individuals representing 19 indigenous yak breeds. Sequencing of DNA samples generated approximately 4.3 terabytes (TB) of raw data, with an average sequencing depth of 8.36×. High-quality sequencing reads were successfully mapped to the reference yak genome, achieving an average alignment rate of 98.4%. Following stringent quality control and filtering, we identified a total of 13.41 million high-confidence single nucleotide polymorphisms (SNPs). These genome-wide SNP markers provide a valuable resource for in-depth investigation into the demographic history and adaptive evolutionary mechanisms of Chinese yak populations, offering critical genomic support for the conservation and innovative utilization of yak genetic resources.

The Yunlong Short-Leg chicken is a local dual-purpose breed in the Dali Bai Autonomous Prefecture of Yunnan Province, China, characterized phenotypically by notably shortened shank and a compact body stature. Nevertheless, the genetic mechanisms responsible for its short-legged phenotype have yet to be elucidated. In this study, Yunlong Short-Leg chickens with different shank lengths-Long-legged and Short-legged phenotypes-were selected as experimental subjects (n = 60, 30 males and 30 females). We performed genome-wide analyses of single nucleotide polymorphisms (SNPs) and insertion/deletion variants (InDels), followed by genome-wide association studies (GWAS) correlating these variants with body weight and morphometric traits. Population genetic analysis indicated no significant differences in genetic background between the Long‑legged and Short‑legged groups, confirming they belong to the same population. GWAS identified shank length as the only trait significantly associated with eight genes on chromosome 7, including IHH, NHEJ1, and MNR2. A comparison of variants on chromosome 7 revealed that the Long‑legged samples exhibited no structural changes, whereas Short-legged samples were categorized as creeper phenotype, exhibiting two non-contiguous deletions and an inversion on chromosome 7: a 13,480bp deletion (positions 22,287,390-22,300,869bp) and an 11,559bp deletion (positions 22,301,448-22,313,006bp), totaling 25,039bp, as well as a 578bp inversion (positions 22,300,870-22,301,447bp). Lethal samples carried a homozygous deletion and the same inversion in this region on chromosome 7. Multiplex PCR with different primers was performed to genotype shank-length phenotypes based on structural genomic variants. Finally, qRT‑PCR analysis confirmed that the relative mRNA expression levels of IHH and NHEJ1 correlated with corresponding locus variation and shank-length phenotypes in Yunlong Short-Leg chicken. These findings provide insight into the genetic basis of the short‑leg phenotype in Yunlong Short-Leg chicken and offer theoretical support for genetic resource conservation and future breeding programs.

Characterizing genomic diversity and population structure is essential for the conservation and sustainable utilization of indigenous sheep genetic resources. Anatolia represents an important center of sheep diversity, harboring multiple populations shaped by diverse breeding histories, ecological conditions, and management practices. In this study, we performed a comprehensive genome-wide assessment of fifteen Anatolian sheep populations using SNP-based data to investigate their genomic diversity, population structure, and genetic relationships. Genetic diversity indices revealed substantial heterogeneity among populations in terms of heterozygosity, allelic richness, minor allele frequency, proportion of polymorphic loci, and inbreeding coefficients, indicating marked differences in within-population genomic variation. Analysis of molecular variance showed that most of the total genetic variation was distributed within populations, although a significant proportion was also attributable to among-population differences. Pairwise FST estimates and principal component analysis demonstrated a structured yet heterogeneous genomic landscape, with some populations exhibiting clear differentiation and others clustering more closely. Phylogenetic analyses further supported this pattern through differences in monophyletic clustering, branch-length dispersion within populations, and patristic distances between populations. Admixture analysis identified three major ancestry components and revealed population-specific differences in ancestry composition and admixture levels. Runs of homozygosity (ROH) analyses showed considerable variation among populations in both the abundance and length distribution of homozygous segments, which was also reflected in class-specific ROH-based genomic inbreeding coefficient (FROH) profiles. Gene flow estimates additionally indicated heterogeneous connectivity across populations. Together, these findings demonstrate that Anatolian sheep populations retain diverse and structured genomic backgrounds despite varying levels of differentiation, admixture, and genomic inbreeding. This study provides a robust genomic framework for understanding the evolutionary relationships among Anatolian sheep populations and offers valuable insights to inform future conservation, management, and breeding strategies.

Comparative transcriptomic analysis of the hypothalamus between tumbler and white king pigeons.

<i>Pseudotsuga sinensis</i> var. <i>brevifolia</i>, formerly known as <i>Pseudotsuga brevifolia</i>, is a native coniferous tree in Vietnam and China. Recently, the number of individuals has decreased significantly and the species continues to be at risk of extinction due to forest fires and logging. This study reported the complete chloroplast (cp) genome of <i>P. sinensis</i> var. <i>brevifolia</i> to establish its genetic relationships with other closely related taxa and to provide a foundation for the future genetic resource conservation of the species. The results revealed that the total length of the cp genome was 122,441bp, with the lengths of the large single-copy, small single-copy, and inverted repeat regions being 66,904 bp, 54,839 bp and 349 bp, respectively. The cp genome of <i>P. sinensis</i> var. <i>brevifolia</i> contains 113 coding genes in total, consisting of 73 protein-coding genes, 36 tRNA genes, and four rRNA genes. The phylogenetic results revealed that <i>P. sinensis</i> var. <i>brevifolia</i> and other <i>Pseudotsuga</i> species are closely related and form a monophyletic group. The results of this study provide additional molecular data for the phylogenetic localization of <i>P. sinensis</i> var. <i>brevifolia</i> and essential data for its identification and resource conservation.

The article presents the results of studies of the allelic pool of breeding bulls of the Kholmogor cattle breed aimed to identify animals for use as donors of semen in genetic resources conservation programs. The sample of breeding bulls (n = 70) was represented by animals born in 1989—2021. 115 831 SNPs that passed through the quality control were selected for analysis, including 73 689 SNPs after LD filtration. Twenty-one of 37 purebred (based on pedigree records) bulls were recognized as genetically purebred animals, based on the membership degree in a Kholmogor-specific cluster of more than 95.0%. The possibility of differentiation of genomic components of the Pechora type of Kholmogor cattle was shown. Six breeding bulls carrying more than 99.9% of the genomic components of this type were identified. The calculation of IBS distances showed a different degree of genetic kinship between donor heifers and breeding bulls: 0.680 to 0.727 with an average value of 0.695 ± 0.001. When developing programs for the conservation of genetic resources to preserve the genomic components of Kholmogor cattle while maintaining the level of biodiversity, it is advisable to use the semen of breeding bulls with confirmed purebredness based on the results of SNP analysis and characterized by the lowest values of genetic kinship estimated by the IBS distances.

Somatic embryogenesis is an artificially induced process that results in the formation of an embryo without the fusion of gametes. Despite its significant potential for plant propagation and conservation of plant genetic resources, many unknowns remain regarding the specific mechanisms and the complexity of genetic, biochemical, and cellular regulations involved. The comparative data on induction of embryogenic tissue and the maturation of somatic embryos, along with additional results, suggest that complex carbohydrate dynamics are among the key factors involved. This review summarizes recent findings focused on early stages of SE, including induction, proliferation, comparison of embryogenic cell lines with contrasting embryogenic capacity, and loss of embryogenic capacity after prolonged cultivation obtained through various methods, including metabolomics, proteomics, transcriptomics, and cytological techniques/histochemical visualization, which highlight the regulation of the energy source starch. This encompasses its synthesis and remobilization, driven by related enzymatic machinery, as crucial for determining embryogenic capacity and prognosis in embryo maturation and germination in many economically important plant species. We also outline potential future directions, critical questions to address, and anticipated advancements in the topic, including the development of new research methodologies.

Due to its high efficiency and safety, oocyte vitrification finds broad application in many fields of life sciences, such as clinical assisted reproduction and conservation of animal genetic resources. However, vitrification may cause cellular damage and reduce the quality of oocytes and their cumulus cells (CCs), which could be closely related to disorders in lipid metabolism. At present, the impact of vitrification upon the lipid profile of oocytes and CCs has not been systematically elucidated. In this study, we used porcine germinal vesicle cumulus-oocyte complexes (COCs) as a model to analyze their lipid characteristics after vitrification and in vitro maturation (IVM), utilizing untargeted lipid metabolomics. Our results showed that an overall count of 37 down-regulated and 8 up-regulated differential lipids was identified in the vitrified oocytes. Pathway analysis confirmed the enrichment in glycerophospholipid metabolism and fat digestion and absorption, etc. Combined with transcriptomic analysis, three enriched pathways were revealed, including the AMPK signaling pathway, metabolic pathways, and fatty acid elongation. On the other hand, a total of four down-regulated and eight up-regulated differential lipids were detected in the vitrified CCs. Pathway enrichment implicated autophagy, glycerophospholipid metabolism, etc. A joint analysis of metabolomic and transcriptomic data revealed four enrichment pathways, including cholesterol metabolism, fat digestion and absorption, regulation of lipolysis in adipocytes, and metabolic pathways. Notably, the supplementation of lysophosphatidylcholine during IVM attenuated oxidative stress, enhanced mitochondrial activity, and enhanced the viability and embryonic development of cryopreserved porcine oocytes. The results indicate that vitrification alters lipids in oocytes and CCs, and the supplementation of lipids plays a role in improving the quality of vitrified oocytes.

Objective: This study evaluated the greasy fleece weight (GFW) and shearing live weight at shearing (SLW) of the Kaçeli sheep breed conserved under Türkiye’s National Animal Genetic Resources Conservation Program.Materials and Methods: The animal material for this study consisted of 291 Kaçeli sheep (283 females and 8 males) raised under semi-intensive conditions on the Çeşme Peninsula. The animals ranged in age from 2 to 7 years. Dirty fleece weights were recorded during shearing in May, and live weights at shearing were measured using digital scales. Due to the small sample size and physiological differences in males, ram data were evaluated only descriptively and were excluded from statistical models. Descriptive statistics, general linear models, least squares means, and phenotypic correlation analyses were performed on the ewe data using SAS software. Farm, age, and live weight were included as fixed effects in the models, and between-group differences were determined using the Duncan Multiple Range Test.Results: Mean GFW was 1.70 kg in rams and 1.26 kg in ewes, while mean SLW was 57.61 kg and 41.40 kg, respectively. Farm effects were significant; Farm 2 showed the highest SLW (45.38 kg). Each 1 kg increase in SLW resulted in a 0.020 kg rise in GFW. Age negatively affected GFW (–0.046) but positively influenced SLW (1.112).Conclusion: Kaçeli sheep display moderate productivity, broad phenotypic diversity, and strong adaptability, providing key data for sustainable breeding and conservation strategies.

Shoot tip cryopreservation is essential for the long-term conservation of plant genetic resources. It provides the only reliable method for establishing a long-term, readily available gene pool of clonally propagated crops and elite in vitro clones used in the pharmaceutical, food, and cosmetic industries. Still, its success is often limited by the inherent sensitivity of many species to the osmotic and chemical stresses imposed by concentrated cryoprotectant (vitrification) solutions and severe dehydration. The optimization of modern cryopreservation protocols primarily focuses on modifying shoot tip preculture, cryoprotectant treatments, or regrowth conditions, while frequently overlooking donor plant preconditioning or relegating it to a secondary role. However, the physiological state of in vitro plants from which apical or axillary shoot tips are extracted may hold the key to successful post-cryopreservation recovery, especially in cryo-sensitive taxa. This review revisits the critical role of donor plant vigor and induced stress tolerance in the cryopreservation of clonal crops by systematically evaluating preconditioning strategies, including cold acclimation, sucrose pretreatment, and the use of growth regulators and signaling molecules such as abscisic, jasmonic, and salicylic acids, involved in stress signaling and tolerance development. The beneficial physiological changes induced by donor plant pretreatment, such as reduced freezable water content and the accumulation of protective compounds, are discussed in the context of contemporary cryopreservation methods. The effects of culture conditions, including the roles of ammonium and nitrates, light quality, culture density and aeration, medium strength, culture age, and subculture duration, are also considered. We analyze how different treatments of in vitro donor plants improve shoot tip tolerance to osmotic and/or chemical toxicity imposed by specific cryopreservation methods to support a material-centered selection of a cryopreservation procedure. Future directions and potential approaches for integrating target donor plant preconditioning into modern cryopreservation protocols for shoot tips, particularly in stress-sensitive species, are discussed.

Exploration and morphological variation of liberoid coffee (Coffea liberica) germplasm as a basis for genetic resource conservation.

This study investigates the genetic diversity, population structure, and adaptive differentiation of Yunnan native cattle (YNC) using whole-genome SNP data from 457 individuals, representing eight cattle populations and two closely related bovine species (Zhongdian yak and Dulong gayal). Genetic diversity analyses revealed a distinct latitudinal gradient from north to south, with the highest diversity observed in the northern Diqing (DQC) and Zhaotong (ZTC) populations. The observed population structure was largely consistent with geographic distribution, identifying distinct ancestral components and complex admixture patterns. Genome-wide selective sweep scans revealed several key candidate genes underlying local adaptation. Notably, GRIA4 and DUOXA2 were associated with cold tolerance in northern populations, and ST3GAL3 and MST1 were implicated in heat stress adaptation in southern populations. Genome-wide balancing selection analyses further detected significant loci, such as MGST1 and SLC36A1, where divergent haplotype frequencies reflected differential selective pressures on milk-related traits between northern and southern populations. Additionally, we detected signals of historical introgression from Zhongdian yak into DQC cattle, highlighting the introgressed gene SLIT3 as a potential candidate associated with high-altitude thermogenesis. Collectively, these results provide a comprehensive genomic framework for the management and conservation of indigenous bovine genetic resources in Southwest China.

Improvement of cryopreserved rooster semen quality by combined application of DMF and ZnO NPs.

High-throughput and cost-effective genotyping technologies are essential for animal genomics and breeding research. In this study, we developed a 100 K single nucleotide polymorphism (SNP) panel for genomic screening of Asian water buffalo using genotyping by target sequencing (GBTS). The panel consists of 101 032 SNPs, selected to include highly polymorphic variants from an existing SNP array-the Axiom 90 K Genotyping Array (16.6%), moderate- to high-impact variants in protein-coding genes (20.3%), swamp- and river-specific variants (10.6%), variants in functional genes associated with economic traits (0.9%), variants located on the X chromosome (0.6%) as well as gap-filling variants (51.0%). To validate the panel, we genotyped 78 buffaloes using the panel as well as whole-genome sequencing (WGS). The panel achieved an average call rate of 99.77%, with a mean genotype concordance of 99.43% across five replicate samples. Population genomics analyses using the panel yielded results highly consistent with those obtained from WGS. When using WGS data as the reference panel, the 100 K SNP panel achieved high imputation accuracy across three buffalo populations-0.89 in dairy river, 0.84 in local river, and 0.81 in swamp buffalo. After applying a stringent quality-control filter (DR2 > 0.9), the retained variants exhibited mean imputation accuracy > 0.90 in all populations while preserving a substantial number of high-quality SNPs (10.40, 5.59, and 2.50 million SNPs, respectively). Overall, the newly designed 100 K SNP panel represents a robust, high-throughput tool for genome-wide genotyping, facilitating the conservation of genetic resources and enabling sustainable genetic improvement of water buffalo populations.

Wild relatives of domestic animals are crucial reservoirs of genetic diversity, yet pervasive hybridization with domestic animals poses significant conservation challenges. Here, we developed a deep learning-based pipeline, consisting of a multi-layer perceptron for SNP panel selection and a Deep & Cross Network for model training, to discern wild relatives from their closely related domestic animals using genomic SNP data. Leveraging the 1960 genomes from 164 red jungle fowl (RJF; Gallus gallus) and 1796 domestic chicken samples, we applied this pipeline to yield the RJF identification model based on a 285-SNP panel. We employed this model to characterize domestic chickens, RJF, and hybrids in the independent genomic datasets from contemporary samples and historical specimens, respectively. The accuracy was 97.8% for historical samples with missing genotypes. The benchmarking multiple hybrid detection tools indicated that the RJF identification model was effective and practical. The further application to the genomic data from wild boar (Sus scrofa), domestic pigs, and their hybrids validated the pipeline. Our method has potential in not only monitoring genetic diversity in wild relatives of domestic animals but also supporting animal genetic resource conservation and management.

Tulipa alberti is an important plant species used for ornamental purposes in Asia, Europe, and North Africa. This study reports the complete chloroplast genome features of this endangered species collected from Kazakhstan using Illumina sequencing technology. The results revealed that the T. alberti chloroplast genome is highly conserved. The genome size is 152,006 bp, comprising a pair of inverted repeats (IR) of 26,330 bp, a large single-copy (LSC) region of 82,169 bp, and a small single-copy (SSC) region of 17,172 bp. The overall GC content is 36.58%. A total of 131 genes were identified, including 85 protein-coding genes, 38 tRNA genes, and 8 rRNA genes. Twenty-eight genes contained introns with lengths ranging from 540 to 2,620 bp. The nucleotide diversity (π) was 0.003257. Analysis of simple sequence repeats (SSR) identified 159 SSR loci in the T. alberti genome, mainly distributed in the LSC region (63.19%), SSC region (18.39%), and IR region (14.37%). Six types of SSR motifs (mono-, di-, tri-, tetra-, penta-, and hexanucleotides) were detected. The most polymorphic protein-coding (CDS) genes were rpoC2, cemA, rbcL, rpl36, psbH, rps3, rpl22, ndhF, ycf1, and matK, which exhibited high sequence variability (SV = 2.581–6.102) and nucleotide diversity (π = 0.004–0.010). Relative synonymous codon usage (RSCU) analysis revealed both preferred and less frequently used codons. The characterization of the T. alberti chloroplast genome provides valuable insights into the conservation of genetic resources, evolutionary and phylogenetic relationships of this rare species. However, to date, data on the complete sequencing and comprehensive bioinformatics analysis of the chloroplast genome of T. alberti remain limited. This study contributes to biodiversity preservation in Kazakhstan and offers a basis for developing effective conservation strategies at the molecular level.

Colostrum is the first mammary secretion produced after parturition and represents a biologically active fluid essential for neonatal survival, immune maturation, and early growth. Despite its importance, breed-related and temporal variations in goat colostrum remain insufficiently characterized, particularly within indigenous production systems. This study comprehensively evaluated the physicochemical, rheological, and microstructural properties of colostrum from four Indonesian goat breeds—Kacang, Local Gorontalo, Saanen crossbreed, and Etawa crossbreed—during the first three days postpartum. Standardized analytical techniques were applied, including proximate composition, amino acid and fatty acid profiling, sodium dodecyl sulfate–polyacrylamide gel electrophoresis, high-performance liquid chromatography, gas chromatography, particle size and zeta potential analysis, rheometry, and confocal laser scanning microscopy. Significant effects of breed and postpartum day were observed. On Day 1, Saanen crossbreeds exhibited the highest crude protein (16.5 ± 0.3%) and fat (6.7 ± 1.5%), whereas indigenous breeds showed higher lactose concentrations. Crossbreeds were enriched in essential amino acids and polyunsaturated fatty acids, including conjugated linoleic acid. Rheological measurements demonstrated higher viscosity (1.95 × 10⁵ Pa·s) and yield stress (418 Pa) in crossbreeds compared with indigenous goats. Confocal microscopy confirmed denser protein–fat matrices in crossbreeds and looser, more heterogeneous microstructures in indigenous breeds. Principal component analysis clearly differentiated breeds and postpartum stages, supporting genotype–phenotype–function relationships. Beyond neonatal nutrition, these results contribute directly to Sustainable Development Goals 2 (Zero Hunger), 3 (Good Health and Well-Being), 12 (Responsible Consumption and Production), and 15 (Life on Land) by improving kid survival, promoting functional dairy innovation, enhancing resource efficiency, and supporting conservation of indigenous genetic resources in sustainable dairy systems.

There is a growing interest in natural bioactive substances, particularly plant-derived secondary metabolites. Polyphenols constitute one of the largest and most significant groups of these metabolites. Rosebay willowherb (Epilobium angustifolium) is well known in traditional medicine and can serve as a reference species for studying its less-known congener, hairy willowherb (E. hirsutum), thereby expanding knowledge of medicinal plants. This study aimed to quantitatively estimate and compare the total phenolic content (TPC) and total flavonoid content (TFC) in the leaves and flowers of Epilobium angustifolium and E. hirsutum, and to identify populations with the highest phytochemical potential. TPC and TFC were quantified using the Folin-Ciocalteu and aluminium chloride (AlCl3) colorimetric assays, respectively, with resulting values regarded as estimates due to the non-specificity of these assays. The results showed that, in terms of TPC, E. angustifolium leaves accumulated 132 ± 3.4 mg_GAE/g (milligrams of gallic acid equivalent per gram of dry plant mass), exceeding those of E. hirsutum by 16.8%; in flowers, the respective values were 153 ± 3 mg_GAE/g, a difference of 1.3%. Regarding TFC, E. angustifolium leaves contained 25 ± 1.4 mg_RE/g (milligrams of rutin equivalent per gram of dry plant mass), which was 20% lower than in E. hirsutum, whereas its flowers accumulated 44 ± 1.4 mg_RE/g, representing a 63% higher content compared with E. hirsutum. The study may contribute to the selection of the Epilobium populations for genetic resource conservation and sustainable utilisation.

Horizontal transfer of mitochondrial DNA into the nuclear genome generates nuclear mitochondrial sequences (NUMTs), which serve as molecular fossils reflecting long-term mitochondrial-nuclear interactions and genome evolution. However, the biological mechanisms governing NUMT integration, retention, and evolutionary fate remain incompletely understood in domesticated animals. Here, using the latest pig reference genome assembly (Sscrofa11.1), we present a comprehensive genome-wide characterization of NUMTs in pigs and provide new insights into their genomic distribution and evolutionary constraints. We identified 513 high-confidence NUMTs, of which 460 were chromosomally mapped, accounting for 0.0106% of the nuclear genome. Beyond increased detection, our analyses reveal that pig NUMTs exhibit non-random origins, preferentially integrate into genomic regions under weak selective constraint, and are frequently associated with repetitive elements, consistent with a DNA repair-mediated insertion mechanism. NUMTs predominantly occur as short, fragmented sequences and show signatures of long-term neutral evolution, while insertions disrupting coding sequences are strongly selected against. Synteny-based analyses further identified clustered NUMT regions and duplicated NUMTs, suggesting secondary genomic duplication events following initial integration. Comparative analysis with the earlier Sscrofa10.2 assembly demonstrates that improved genome quality substantially enhances NUMT detection, particularly in repetitive and GC-rich regions, clarifying previously ambiguous sequence-context associations. Together, this high-quality pig NUMT map provides a robust foundation for future functional, evolutionary, and population-level investigations and contributes to the conservation and utilization of pig genetic resources.

In vitro culture of callus is an effective method for conserving the genetic resources of economically important crops. However, continuous subculturing is a costly and labor-intensive process. Therefore, establishing an efficient in vitro long-term conservation system applicable to various plant species is required. In this study, calli derived from five medicinal plant species, Camellia japonica (Cj), Centella asiatica (Ca), Ligusticum afficinale (Lo), Panax ginseng (Pg), and Sageratia thea (St) were used to optimize storage conditions and establish a suitable in vitro conservation strategy. Calli cultures were maintained on the appropriate culture medium at 5 °C for 120 days. Cell viability and regrowth rate were assessed during the storage period, and correlations between growth and antioxidant traits were examined. Subsequently, pretreatment optimization using sucrose (3–9%), MeJA (0-200 µM), and CTR (0–20 mg/L) was performed using RSM, and the effects of pretreatment and storage temperature on callus conservation were evaluated. In addition, machine learning models such as GRNN, MLP, RF, SVR, and XGBoost were applied to the experimental data. The findings demonstrated that, in comparison to Ca and St, Lo, Pg, and Cj exhibited noticeably higher antioxidant capacity while maintaining high cell viability and regrowth rates. Interestingly, Ca and St drastically decreased viability and regrowth after 60 days, whereas Lo, Pg, and Cj maintained viability and regeneration for up to 90 days. Both TPC and AC (measured by FRAP assay) showed a high positive correlation with cell viability and growth rate, according to correlation analysis. RSM predicted that the optimal pretreatment medium for enhancing antioxidant capacity was the species-specific proliferation medium supplemented with 3% sucrose, 135 µM MeJA, and 20 mg/L CTR, while the highest TSSC was achieved using the species-specific proliferation medium supplemented with 9% sucrose and 200 µM MeJA. When the storage temperature was set to 15 °C following the antioxidant-enhancing pretreatment derived from the RSM optimization, all five species showed improved cell viability and regrowth rates, among the storage methods. Among the ML models tested, XGBoost demonstrated the most stable and accurate predictive performance for both viability and regrowth during in vitro conservation. SHAP-based analysis of the XGBoost model, focusing on regrowth rate, revealed that storage duration was the most influential factor affecting regrowth prediction, followed by storage temperature, while pretreatment conditions showed secondary but meaningful contributions. This study demonstrates that long-term callus conservation is closely associated with AC and TPC. Medium supplemented with sucrose 3%, 135 µM MeJA, and 20 mg/L CTR, followed by storage at 15 °C, significantly improved viability and regrowth, and calli could be maintained up to 120 days without subculturing. This approach provides an efficient and broadly applicable in vitro strategy for the conservation of diverse plant genetic resources.