Over the past two decades, omics and big data have shifted plant molecular biology from single-gene, hypothesis-driven studies to systems-level, data-driven discovery. As datasets expand in scale and diversity, bioinformatics software has become essential for routine analysis and interpretation. However, the efficiency of data exploration and evidence integration has not kept pace with data growth, leaving many datasets underutilized and only slowly translated into biological insight. A central bottleneck is the widening gap between the limited data analysis skills of many experimental biologists and the increasing complexity of biological data. TBtools was developed to narrow this gap by providing low-barrier, interactive functions for common plant omics tasks, and it has been broadly adopted. Here, we use TBtools as a decade-long case study to discuss why certain local tools achieve broad adoption in plant omics research, distill eight actionable design recommendations, and propose four capacity pillars for next-generation local workbenches: project-level data management, reproducible workflow construction, elastic remote computing, and AI-assisted navigation and automation. Together, these lessons provide a practical roadmap for accelerating the translation of omics data into biological insights.

Durian (Durio zibethinus) is an economically important tropical fruit endemic to Southeast Asia. Hainan is a new region for durian cultivation. Foliar diseases of durian were common at all durian orchards during field surveys carried out from October 2023 to October 2024 in Hainan, China. In this study, Fusarium species associated with durian foliar disease in Hainan (China) were studied based on multilocus phylogenetic analyses using translation elongation factor 1-α (tef1), partial RNA polymerase second largest subunit (RPB2), and calmodulin (CaM). Six Fusarium species, F. pernambucanum, F. sulawesiense, F. hainanense, F. irregulare, F. mangiferae, and F. concentricum, were identified. Pathogenicity tests showed that all six species were pathogenic to detached wounded and unwounded durian leaves except F. irregulare, which only induced visible symptoms on wounded durian leaves. The findings from this study expand the pathogenic fungal species on durian as this is the first report of these Fusarium spp. causing durian leaf disease worldwide.

Fruit shape is a commercially important trait in litchi, yet the genetic mechanisms that regulate its development remain largely unresolved. OVATE Family Proteins (OFPs) are plant-specific transcriptional regulators known to play key roles in fruit morphology, organ growth, and overall plant development. In this study, we identified 14 OFP genes distributed across eight chromosomes in the litchi genome. Phylogenetic reconstruction and conserved motif analyses demonstrated that all LcOFPs contain the characteristic OVATE domain and can be grouped into three subfamilies based on motif composition. Promoter cis-element analysis further suggested that LcOFPs may respond to diverse phytohormone signals and abiotic stresses, indicating their involvement in multiple regulatory pathways. Integrated RNA-seq and RT-qPCR analyses revealed that LcOFP3, LcOFP6, LcOFP10, LcOFP13, and LcOFP14 show tissue-specific expression patterns and dynamic transcriptional changes during fruit development in two cultivars with contrasting fruit sizes. Heterologous overexpression of LcOFP3 significantly modified fruit morphology in tomato. These findings clarify the genetic mechanisms through which OFP genes regulate fruit shape in litchi and highlight promising genetic targets for future litchi breeding and improvement.

While mutagenesis breeding has been widely applied in major crops, its implementation in tropical horticultural species remains limited. In this study, we established a pingyangmycin-induced mutagenesis system using in vitro–cultured embryogenic callus of litchi. Treatment with gradient concentrations of pingyangmycin (0–40 mg/L) revealed an inverse relationship between chemical dosage and cell viability, with somatic embryogenesis completely inhibited at 40 mg/L. Whole-genome resequencing of treated callus identified 307,629 high quality somatic mutations (255,721 SNVs and 51,908 indels), with the 20 mg/L treatment group exhibiting the highest number of mutations. Further resequencing of 40 regenerated mutant lines from the cultivars ‘Feizixiao’ and ‘Lingnan15’ revealed 1,703,663 and 1,281,489 somatic variants, respectively, corresponding to mutation frequencies of 1.8×10-4 per site and 1.4×10-4 per site, both markedly exceeding typical EMS-induced rates in crops. Transition mutations predominated, accounting for 77% of total single-nucleotide substitutions. Short indels (<3 bp) comprised over 70% of all insertion–deletion events, indicating pingyangmycin’s propensity to induce base substitutions and small indels via oxidative and double-strand break repair pathways. The regenerated mutant plants exhibited substantial leaf morphological variation, confirming effective mutagenic diversification. Collectively, this study provides the first evidence of pingyangmycin’s mutagenic efficacy in litchi, establishes a reliable system for mutant library construction, and delivers valuable genetic resources for functional genomics and molecular breeding applications.

Somatic embryogenesis (SE) is an important factor influencing the in vitro regeneration efficiency in litchi (Litchi chinensis Sonn.). Although putrescine (Put) and D-arginine (D-Arg) supplementation can regulate embryonic callus (EC) proliferation and somatic embryo induction in litchi, the underlying mechanism remains unclear. This study aimed to investigate the effects of exogenous Put and D-Arg on the histomorphological changes, polyamine (PA) contents, and PA metabolism-associated enzyme activities of 'Feizixiao' litchi EC. ECs were cultured on original proliferation medium (M3) and M3 supplemented with 15.0 mg·L−1 Put (P3) or 0.35 g·L−1 D-Arg (Ar3) for three weeks. Exogenous Put and D-Arg significantly increased the EC proliferation rate. Put reduced the numbers of induced somatic embryos and regenerated plantlets, while D-Arg had the opposite effects. Histomorphological analysis showed that exogenous Put or D-Arg inhibited proembryo differentiation during EC proliferation. On the M3 medium, EC cells first divided into two cells and then sequentially differentiated through the multicellular proembryo, globular, heart-shaped, and cotyledonary embryo stages. On P3 and Ar3 media, embryonic cells enlarge more significantly, underwent repeated cell divisions, and subsequently formed a meristematic mass from which dicotyledonous embryos initiated. P3 and Ar3 media both enhanced the endogenous PA contents and decreased the endogenous ornithine decarboxylase (ODC) and polyamine oxidase (PAO) activities. Exogenous Put inhibited the endogenous arginine decarboxylase (ADC) activity and enhanced the diamine oxidase (DAO) activity, while exogenous D-Arg had the opposite impact. In further experiments, ECs from P3 and Ar3 medium were subsequently cultured on M3, P3 and Ar3 media, respectively. In the control, ECs from M3 medium were cultured on M3 (M3M3) medium. ECs exhibited the highest proliferation rate on P3Ar3 medium which was higher than that of M3M3, while the EC proliferation rates of the remaining treatments were lower than that of M3M3. The Ar3Ar3 medium produced the highest numbers of somatic embryos, dicotyledonous embryos, and regenerated plantlets, while the P3P3 medium yielded the fewest somatic embryos and plantlets. Compared with the control, the Put content decreased and the spermine (Spm) content increased in ECs in all other treatments. In addition, the total PA content also increased, except for in the P3Ar3 treatment. The ODC enzyme activities in all treatments were lower than that of the control, while enhanced ADC, DAO, and PAO activities were found in the P3P3, Ar3P3, and Ar3Ar3 treatments, respectively. These results suggest that exogenous Put or D-Arg can stimulate endogenous Put synthesis via regulating enzyme activities and influencing EC proliferation and SE in litchi.

2,4-Dichlorophenoxyacetic acid (2,4-D) is a vital exogenous auxin for the induction and proliferation of litchi embryogenic callus. At present, its molecular regulation mechanism remains unclear. In this study, transcriptome sequencing samples were selected based on different cell growth phenotypes observed in 'Feizixiao' litchi embryogenic callus cultured in liquid medium with or without 2,4-D. By integrating transcriptome profiling with weighted gene co-expression network analysis (WGCNA), we identified key genes and signaling pathways dynamically responsive to 2,4-D concentration changes. We identified 558 commonly differentially expressed genes (DEGs), of which 117 were up-regulated and 387 were down-regulated; functional enrichment analysis revealed significant enrichment in the "plant hormone signal transduction" and "phenylpropanoid biosynthesis" pathways. In the former pathway, genes such as AUX28, GH3.17, GH3.6, and ARR5 were up-regulated; in the latter, by comparison, beta-glucosidase 47 and Peroxidase 61 exhibited increased expression levels induced by 2,4-D. Furthermore, among these DEGs, 57 transcription factors belonged to 24 families. Notably, VRN1, FEZ, and DOF5.4 were significantly and rapidly induced by 2,4-D. WGCNA results demonstrated a significant positive correlation between the yellow module and 2,4-D treatment. Small heat shock protein (sHSP) genes constituted the core hub genes in the yellow module. Through Venn analysis of DEGs and key modules, 38 cross-genes were identified, of which non-specific lipid-transfer protein-like genes (nsLTP) were found to be specifically up-regulated without 2,4-D. The transcription factors and genes identified work in synergy to ensure the formation and sustained proliferation of embryogenic callus by precisely regulating the dynamic balance of auxin and cytokinin within cells and maintaining the stability of cell structure. Our findings provide a crucial theoretical foundation for understanding the molecular mechanism of 2,4-D in regulating litchi embryogenic callus proliferation.

[Objective]Litchi(Litchi chinensis Sonn.)is an economically valuable fruit species in southern China.Breeding new variety is very important for meeting consumer needs and improving eco-nomic efficiency.However,the species'allogamous reproductive strategy drives extreme genomic het-erozygosity,presenting formidable barriers to precision breeding and genetic manipulation.To address these constraints,cell suspension culture emerges as a transformative biotechnological platform,offer-ing dual strategic capacities.It would generate homogeneous cell populations essential for protoplast isolation,somatic hybridization,and transgenic development;Moreover,it would enable industrial-scale biosynthesis of high-value phytochemicals.This study systematically optimized critical parame-ters governing litchi anther-derived callus growth in suspension systems,including macro-/micronutri-ent formulations(inositol,2,4-D,sucrose),organic growth modulators[lactalbumin hydrolysate(LH),coconut water(CW)],and inoculation density thresholds.The multifactorial optimization strategy aimed to establish a robust,scalable suspension culture protocol yielding high-quality embryogenic cul-tures,thereby creating foundational biomaterials for advanced applications in litchi protoplast technolo-gy and cross-species genetic engineering initiatives.[Methods]Anther-derived embryogenic callus of Feizixiao litchi was used as the initial material.Suspension cultures were established in liquid MS medi-um under dark conditions(25±2℃,120 r·min-1).A series of experiments were conducted to optimize culture conditions:Inositol(0-0.2 g·L-1),2,4-D(0.1-2 mg·L-1),sucrose(10-40 g·L-1),LH(0-0.4 g·L-1),CW(0-200 mL·L-1),and inoculum densities(10-40 g·L-1)were tested for their effects on cell prolifera-tion,single-cell viability,and biomass.Suspension cultures were subculture every 4 days,filtered through a 0.85 mm sieve,and monitored for growth parameters[fresh weight(FW),dry weight(DW),packed cell volume(PCV),pH,conductivity).Embryogenic callus proliferation,somatic embryo differ-entiation,maturation,and plant regeneration were induced using specific solid media supplemented with growth regulators(NAA,ZT,ABA)and high sucrose concentrations.[Results]Single-factor trial results showed that the effects of inositol,2,4-D,sucrose,CW and initial inoculum density on the callus morphology(single cell,little cell group and density)and proliferation rate(FW,DW)were significant(P＜0.05),while the effect of LH was not significant.The embryogenic suspension system of litchi was established.The optimal treatment regimen was as follows:Anther embryogenic callus of Feizixiao li-tchi was inoculated into liquid medium containing MS+inositol 0.15 g·L-1+2,4-D 1 mg·L-1+sugar 20 g·L-1+CW 50 mL·L-1.The initial inoculum density was 1.5 g per 50 mL of culture medium.The suspension culture was maintained under dark conditions at(25±2)℃ with 120 rpm orbital shaking.The subculturing was performed every 4 days for 3-4 cycles,followed by filtration through 0.85 mm mesh,after that the subculturing interval was extended to 7 days.After approximately 20 days of total cultivation,an optimal suspension cell line was obtained,characterized by predominantly round or near-round cell morphology,excellent dispersion and homogeneity,rapid cell division and growth,abundant and active cytoplasmic structures,and a clear,transparent suspension.The growth parameters of the li-tchi suspension culture(single-cell yield,biomass accumulation,and cell packed volume)followed a sigmoidal("S"-shaped)growth curve.The culture progression was divided into three phases:lag phase(days 1-2):minimal single-cell dispersion,slow increases in FW,DW and packed cell volume(PCV),logarithmic growth phase(days 3-6):rapid cell division with significant increases in single-cell count,FW,DW,and cell density,and stationary phase(days 7-16):stabilized cell growth with gradual increases in FW,DW and PCV.Beyond this phase,the prolonged culture led to cell senescence and fragmentation,accompanied by sharp declines in viable cell count and survival rate,with minimal increases in FW,DW and PCV.The pH of the suspension initially showed a slight increase followed by continuous decline throughout the culture period.The electrical conductivity exhibited a consistent downward trend during the entire suspension culture process.[Conclusion]An optimized embryogenic suspension system was successfully established for Feizixiao litchi using MS medium supplemented with inositol 0.15 g·L-1,2,4-D 1 mg·L-1,sucrose 20 g·L-1,and CW 50 mL·L-1.This system exhibited high uniformity,rapid pro-liferation,and stable embryogenic potential,enabling efficient somatic embryo production and plant re-generation.The findings could advance litchi biotechnology applications,including mutagenesis,proto-plast isolation,and synthetic seed production,and offer a reference for recalcitrant woody species.

Litchi chinensis var. fulvosus is an important wild litchi resource in Yunnan, China, valued for favorable agronomic traits such as early flowering, early ripening, multiple flowering cycles, and high fruit-setting ability. However, its genetic diversity and population structure remain poorly understood. In this study, 192 accessions were collected from ten counties in Yunnan Province to evaluate their geographic distribution, leaf phenotypic variation, molecular diversity, population structure, and core collection composition. Eight descriptive leaf traits, nine quantitative leaf traits, and ISSR genotyping data from seven primers were analyzed. The accessions were distributed across an altitudinal range of 169-1470 m, with clear habitat differentiation among trees of different ages. Morphological analysis revealed substantial leaf variation, with mean diversity indices of 1.19 for descriptive traits and 2.76 for quantitative traits. ISSR analysis generated 49 scorable bands, of which 34 were polymorphic, corresponding to a polymorphism rate of 68.45%. The mean Shannon-Wiener diversity index was 0.3101, indicating detectable but relatively limited molecular diversity. Integrated phenotypic and molecular analyses divided the germplasm into two subpopulations. A core collection comprising 30 accessions (about 15% of the initial population) showed the best balance between sampling efficiency and diversity retention. These results provide a practical basis for the conservation, evaluation, and efficient utilization of L. chinensis var. fulvosus genetic resources and will support breeding and genetic improvement of litchi.

Pericarp dehydration is a primary factor contributing to litchi fruit browning, severely limiting the marketability of the product. Aquaporins (AQPs), a class of multifunctional membrane proteins, play a pivotal role in regulating the transmembrane transport of water in plants. However, the specific involvement of AQPs in the dehydration and browning of litchi fruit, along with their underlying regulatory mechanisms, remain to be fully elucidated. The results of the present study demonstrated that control (unpackaged) litchi fruit exhibited complete browning, which was accompanied by severe weight loss (10.2 %), discoloration, and membrane damage after 60 h of storage. These rapid physiological changes were alleviated by the use of polyethylene bag packaging, an effective high-humidity maintenance method. Transcriptomic analysis revealed differential expression of 12 AQP genes across the experimental groups, among which four genes (LcPIP1;1, LcPIP2;2, LcPIP2;5 and LcSIP2;1) were upregulated, while eight genes (LcPIP2;7, LcTIP1;2, LcTIP2;1, LcTIP4;1, LcTIP1;1, LcSIP2;2, LcNIP1;1 and LcNIP1;2) were downregulated in control fruit during storage. The changes in the expression of most AQP genes observed in control fruit were substantially counteracted by packaging treatment, thereby ameliorating pericarp dehydration and browning in litchi fruit. Furthermore, LcMYB6, a MYB transcription factor that is induced by packaging treatment, binds to the promoters of LcPIP1;1 and LcPIP2;5. Promoter binding assays verified that LcMYB6 is a nuclear protein that directly represses the transcription of LcPIP1;1 and LcPIP2;5. Transient silencing of LcMYB6 promoted the upregulation of LcPIP1;1 and LcPIP2;5 expression while accelerating pericarp dehydration and browning. These results indicate that LcMYB6 may increase litchi dehydration resistance by directly repressing the transcription of AQP genes (LcPIP1;1 and LcPIP2;5), potentially offering new strategies for controlling pericarp browning in litchi fruit.

Durian ( Durio zibethinus Murray), the king of fruits, is an edible and economically important tropical fruit endemic to Southeast Asia. Durian is affected by Colletotrichum, which is one of the most important genera of plant pathogenic fungi, especially on tropical and subtropical crops. In this study, Colletotrichum species associated with durian in Hainan (China) which was a new region for durian cultivation were studied using phylogenetic and morphological analyses. The results of molecular identification based on internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase ( GAPDH ), beta-tubulin ( TUB2 ) and the mating type locus MAT1-2 ( ApMat ) along with microscopic identification indicated that seven species from three species complexes were identified, including C. fructicola, C. siamense, C. queenslandicum and C. endophyticum from the gloeosporioides species complex; C.plurivorum and C. musicola from the orchidearum species complex, and C. gigasporum from the gigasporum species complex. Colletotrichum fructicola and C. siamense were the main Colletotrichum species associated with durian in Hainan. Pathogenicity tests showed that all seven species could infect durian leaves using a wound inoculation method but only C. fructicola, C. queenslandicum and C. endophyticum could infect using a non-wound inoculation method. The findings from this study enhance the knowledge of durian diseases and lay an essential foundation for devising effective disease management approaches. Further largescale surveys are necessary, including sampling different sites and plant tissues, pathogenicity testing on a wider range of host genotypes and fungicide-sensitivity evaluation among and within the different Colletotrichum species collected.

Litchi (Litchi chinensis Sonn.), an important fruit tree in tropical and subtropical regions, possesses substantial economic value. The branch- and leaf-related traits of litchi have a significant impact on litchi yield and quality. However, due to limitations such as the density of the genetic linkage map, there have been few studies on mapping QTLs of branch- and leaf-related traits. In this study, a high-density genetic map was constructed by next-generation sequencing (NGS) using an F1 population of 264 progenies, derived from the cross between the cultivars ‘Sanyuehong’ and ‘Ziniangxi’. A total of 2 574 high-quality BINs (binomial intervals) were obtained, and a genetic linkage map was constructed with a total length of 1 753.3 cM and an average marker distance of 0.68 cM. With the genetic map and the phenotyping of single leaf length (SLL), single leaf width (SLW), leaf shape index (LSI), weight of specific leaf (WSL), petiole length (PL) and compound leaf length (CLL) measured in three seasons, 11, 9, 9, 10, 9 and 12 QTLs were detected for SLL, SLW, WSL, LSI, PL and CLL traits, respectively. Among these QTLs, five QTLs were consistently detected in two seasons and 12 pleiotropic QTLs were identified for at least two traits. These findings will provide new insights for the gene cloning for branch- and leaf-related traits as well as marker-assisted selection (MAS).

Seedless litchi (Litchi chinensis ‘Wu He’) exhibits an excellent parthenocarpic trait, the physiological regulation of which during early fruit development is critical for ultimate yield. Sugar availability serves as the predominant factor governing both fruit abscission and development. However, the molecular mechanisms by which sugar signaling mediates hormonal networks to regulate these processes in seedless litchi remain unclear. This study explored the regulatory patterns in seedless litchi during the massive abscission stage (MA stage) and subsequent developmental stages (SD stages), through integrated physiological and transcriptomic analyses. At the MA stage (15 days after flowering, 15 DAF), the concentrations of glucose and fructose in fruits dropped to their lowest levels. Concurrently, auxin and abscisic acid (ABA) levels increased significantly, while the ethylene (ET) precursor (1-aminocyclopropane-1-carboxylic acid, ACC) content declined. Tissue section revealed that both cell size and cell number were at their lowest, indicating delayed fruit development. Upon entering the SD stages (22 DAF, 29 DAF, 36 DAF), fruit abscission progressively decreased as sugar availability improved. During these stages, cytokinin (iP-type-CK) and gibberellin (GA3) levels increased, accompanied by synchronized increases in cell volume and number, leading to continuous fruit enlargement. Transcriptomic analysis demonstrated that differentially expressed genes were primarily enriched in pathways related to starch and sucrose metabolism, hormone metabolism and signal transduction. During the MA stage, expression of genes such as MAT, ACS, ACO, CTR1, ZEP,NCED, PP2C, and SnRK2 was upregulated, whereas EIN2 was downregulated. In contrast, the SD stages exhibited upregulation of IPT,B-ARR, KAO, GID1, GID2, TFs and DELLA genes, alongside downregulation of CKX, A-ARR, and GA2ox. These gene expression patterns were consistent with fluctuations in hormone levels and collectively regulated fruit abscission and development. RT-qPCR validation confirmed the accuracy of the RNA-Seq results. Deficiencies in fructose and glucose mediate ABA and ET accumulation while disrupting polar auxin transport, thereby exacerbating fruit abscission. Following massive fruit abscission, improved sugar availability enhanced the effects of CK and GA, which promoted cell division and expansion, resulting in gradual fruit enlargement. These findings elucidate the sugar signaling-mediated hormonal regulatory networks during seedless litchi abscission and development, providing a theoretical foundation for achieving high and stable yields in orchards.

Litchi downy blight is a destructive disease that severely reduces yield and causes postharvest losses in litchi fruit. The varieties ‘Heiye’ (HY) and ‘Guiwei’ (GW) were identified as highly resistant and susceptible to Peronophythora litchii infection, respectively. We further investigated the differences between resistant and susceptible varieties through transcriptomic and biochemical analyses to uncover the mechanisms underlying postharvest resistance to litchi downy blight in litchi fruit. RNA-seq analysis of postharvest litchi fruit infected with P. litchii revealed that differentially expressed genes between HY and GW were primarily associated with phenylpropanoid biosynthesis and reactive oxygen species (ROS)-related pathways. The analysis of key metabolites in phenylpropanoid biosynthesis showed a significant difference in lignin content between the two varieties. Fifteen upregulated genes (C4H-1, COMT-1, CAD-6, and POD-7) were identified by RNA-seq and qRT-PCR. These genes correspond to highly active enzymes of C4H, CAD and POD involved in lignin biosynthesis, which were detected in P. litchii-inoculated HY fruit compared to GW. These factors contributed to the higher lignin content in HY fruit. Additionally, the ROS scavenging ability of HY pericarp was superior to that of GW, as evidenced by reduced ROS and malondialdehyde (MDA) levels, increased activities of superoxide dismutase (SOD) and catalase (CAT), and increased expression of two SOD genes and two CAT genes following P. litchii infection. These findings suggest that lignin biosynthesis and ROS metabolism play crucial roles in maintaining litchi fruit resistance to P. litchii, providing valuable insights for breeding and developing resistant varieties.

Pericarp browning of postharvest litchi is a significant obstacle to the industry's high-quality development. Water loss from the pericarp is a key factor triggering browning, but the regulatory mechanism of water metabolism and its relationship with browning remain unclear. In this study, we found that aquaporin activity inhibitors (HgCl2) can delay both water loss and browning in litchi. LcPIP2;4, a PIP family member exhibiting high expression in the litchi pericarp and the greatest water transport activity, is significantly downregulated during water loss and browning. Further analysis revealed that HgCl₂ suppresses both the expression and water transport activity of LcPIP2;4, indicating a close association with the observed browning phenotype. By constructing transient overexpression fruits and transgenic callus tissues of LcPIP2;4 and measuring the water loss rate and browning index, we confirmed that LcPIP2;4 positively regulates water loss and browning in litchi. Through weighted gene co-expression network, LcPIP2;4 promoter sequence and qRT-PCR analysis, we identified 10 potential interacting transcription factors. Yeast one-hybrid, dual-luciferase reporter assay, chromatin immunoprecipitation analysis and electrophoretic mobility shift assay confirmed that LcMYB306 specifically binds to the LcPIP2;4 promoter. In LcMYB306 overexpressing fruits and embryogenic callus, LcPIP2;4 expression was suppressed, resulting in delayed water loss and browning. In contrast, in CRISPR/Cas9-edited LcMYB306 callus, LcPIP2;4 expression was upregulated, and water loss and browning were accelerated, confirming that LcMYB306 negatively regulates this process. This study demonstrates that LcMYB306 delays postharvest water loss and browning in litchi by repressing LcPIP2;4 transcriptionally expression. It provides a theoretical foundation and key target gene for developing litchi varieties resistant to browning.

Litchi, an important tropical fruit, is severely affected by anthracnose disease. However, the mechanism of its disease resistance response remains unknown, and resistant accession genetic resources and resistance-related genes have not yet been identified. In this study, 82 accessions of litchi were evaluated for resistance to Colletotrichum gloeosporioides, and the accessions ‘Haiken 5’ and ‘Nongmei 5 hao’ were identified as resistant and susceptible, respectively. Leaves from these two accessions were inoculated with C. gloeosporioides and collected at 6 and 24 h for use as materials for transcriptome analysis. Analyses of the differentially expressed genes (DEGs) between the accessions and their controls, which were inoculated with potato dextrose agar medium, revealed that the resistant accession presented more DEGs with smaller changes in magnitude, whereas the susceptible accession presented fewer DEGs with greater changes in magnitude. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed, and phenylpropanoid biosynthesis, amino sugar and nucleotide sugar metabolism, and plant–pathogen interactions were identified as common pathways. Chitinase activity, oxidoreductase activity, aminoglycan and glucosamine-containing compounds, and cell wall metabolic processes also participated in the defence reaction. Salicylic acid signalling in litchi leaves contributed to resistance to C. gloeosporioides. Short Time-series Expression Miner (STEM) and weighted correlation network analysis (WGCNA) were also employed to evaluate the gene expression trends and identify highly correlated genes. Litchi accessions presented different resistance responses to anthracnose disease. Small changes in the expression levels of critical resistance-related genes were sufficient to produce the defence reaction. Calcium ion regulatory mechanisms and transcription factors have been preliminarily identified as contributors to disease resistance. Multiple pathways and molecular processes participate in the defence response. These results identify candidate genes and pathways involved in litchi plant defence against anthracnose.

The efficiency of in vitro regeneration in litchi (Litchi chinensis Sonn.) is highly influenced by the type and concentration of plant growth regulators (PGRs), particularly auxins. This study evaluated the effects of phenylacetic acid (PAA) and its derivatives—4-chlorophenylacetic acid (CPA) and 4-iodophenylacetic acid (IPA)—on callus proliferation, somatic embryogenesis, and plantlet regeneration in ‘Feizixiao’ litchi, as well as to establish an efficient regeneration protocol. The inclusion of CPA or IPA in callus proliferation medium significantly enhanced the proliferation rate, with 20 mg·L−1 CPA being the most effective. The highest number of somatic embryos per gram of fresh embryonic callus weight (gFW−1) (1131 embryos·gFW−1) was observed with 40 mg·L−1 PAA in proliferation medium. The addition of 10 mg·L−1 IPA to the proliferation medium yielded the highest plantlet regeneration rate (50 plantlets·gFW−1). Supplementing the somatic embryo induction medium with 5 mg·L−1 PAA resulted in 460 somatic embryos·gFW−1 and 86 regenerated plantlets·gFW−1. These findings indicate that PAA and its derivatives are effective PGRs for the in vitro regeneration of litchi, providing a valuable protocol for the propagation of elite cultivars.

Genetic breeding and molecular identification in varieties depend on high-performance genotyping tools. The high heterozygosity of the litchi genome contributes to increased resequencing costs and elevated error rates in hybridization-based genotyping methods. In this study, a liquid chip named Litchi40K v1.0 was developed with high-depth resequencing data from 875 litchi samples, and its efficacy was validated across three different populations. In the L. chinensis var. fulvosus population, three subpopulations characterized by spatial distribution, and a total of 1110 genes were identified in the genomic regions with subpopulation differentiation. Additionally, a total of 30 significant signals associated with diverse agronomic traits were identified. The H002 haplotype of LITCHI02696, dominant in the Sub2 subgroup, significantly increased the soluble solid content in the L. chinensis var. fulvosus population. In a hybrid F1 population, a high-density genetic map was constructed and 79 dwarfing-related QTLs were identified with the liquid chip. An NAC transcription factor was identified as a candidate gene with a heterozygous frameshift variant in the male parent. To facilitate the digitization of germplasm resources, 384 SNPs were selected, and the DNA fingerprint map revealed clear genetic relationships and a total of 10 potential synonym groups or instances of bud mutations were identified in 164 main cultivated litchi varieties. This study provides cost-effective, flexible, and versatile liquid chip for genetic analysis and digitalization of germplasm resources in litchi.

Chloroplast and mitochondrial genomes, which coexist with the nuclear genome, are extensively used in gene function studies, evolutionary analyses, and targeted breeding. However, assembling these organelle genomes remains challenging due to frequent recombination events and abundant repetitive sequences. Although some assembly tools are available, many are difficult to install or require extensive parameter tuning and computational resources. To address these limitations, we present a high-fidelity, data-driven mitochondrial genome assembly toolkit (HiMT), a user-friendly, out-of-the-box software solution that enables one-click chloroplast and mitochondrial genome assembly using default parameters, particularly for plant species. HiMT automatically estimates read coverage depth and employs a fixed k-mer prefix strategy to minimize computational demands, making it suitable for use on standard laptops. Benchmark tests show that HiMT delivers complete assemblies with a high success rate, fast runtimes, and low hardware requirements. Additionally, HiMT features a graphical user interface (GUI) and generates interactive reports for assessing the quality of organelle genome assemblies. We anticipate that HiMT will facilitate high-quality plant mitochondrial genome research and significantly streamline organelle genome assembly workflows. To support the research community, HiMT is freely available to non-commercial users at https://github.com/tang-shuyuan/HiMT.

Postharvest pericarp browning, caused primarily by the enzymatic oxidation of phenols, reduces the shelf life and market value of litchi fruit and is considered a major limitation for the development of the litchi industry. Previous studies have shown that polyphenol oxidase (PPO) is a key enzyme and that flavonoids are important substrates for enzymatic browning; however, direct evidence is still lacking. This study investigated the differences in the browning process among the wild type (WT) and four PPO gene-edited litchi calli to verify the function of PPO in the browning of litchi tissues. Compared to the WT callus, the proliferation rate, relative expression of litchi PPO gene (LcPPO), PPO activity and color changes significantly decreased or slowed down in all gene-edited calli, indicating that the latter exhibited a slower browning process. Using a liquid chromatography tandem mass spectrometry approach (LC-MS/MS), 83 metabolites of flavonoids were identified, of which 58 were differentially accumulated metabolites (DAMs). Venn analysis revealed 12 common DAMs across different genotypic contrasts that were mostly enriched in the flavonoid biosynthesis pathway. It was presumed that the decrease of LcPPO expression in gene-edited calli led to the reduced PPO activity, then reduced the (-)-epicatechin oxidation. The accumulation of (-)-epicatechin caused the common upregulation of procyanidin B2 and upstream substances such as dihydrokaempferol, taxifolin, naringenin chalcone, 7,4'-dihydroxyflavone, and rutin in their biosynthesis pathways. The results provide novel evidence that (-)-epicatechin acts as the primary direct substrate in the enzymatic browning reaction mediated by PPO.