Comparative metabolomics and transcriptome analysis provide insights into branching changes induced by heterografting in Pinus massoniana seedlings.

The investigation of dwarfing rootstocks for the establishment of high-generation seed orchards is a prospective avenue of research. In this investigation, Pinus massoniana, Pinus yunnanensis var. pygmaea (P. pygmaea), and P. elliottii seedlings were used as rootstocks for grafting with P. massoniana scions. Grafting P. massoniana onto P. pygmaea rootstock resulted in observable phenotypic alterations in lateral branches, apical buds, and needle length. Certain characteristic metabolites of rootstocks, such as fatty acyls, pregnenolones, steroids, and steroid derivatives, were found to be highly expressed in scions after grafting. RNA-seq analysis revealed MYB-related, SBP, and bHLH demonstrating a significant positive correlation, while C2H2 and Orphans exhibited negative correlations with the differential intensity of metabolites related to lipids and lipid-like molecules. This study offers valuable insights for the establishment of rootstock breeding programs.

Metabolite components and nutritional composition of the endosperm in seven species from Gleditsia.

As an important agricultural product, the endosperm portion of Gleditsia sinensis seeds, called "zào jiǎo mǐ" (ZJM) in Chinese, has gradually gained popularity and has been accepted by the public. However, there is limited information on the nutritional value and metabolic components of endosperm among Gleditsia. This study compared the endosperm composition among seven species. The types of metabolites, content of nutrients and amino acids were determined. A total of 4495 types of metabolites were detected. Galactose metabolism (gmx00052) was enriched in all combinations compared with G. sinensis. The polysaccharides content ranged from 51.49 to 80.37 g/100 g. Based on considerations of growth rate, seed yield, amino acid content, and interspecific differences, G. fera could be an alternative planting option to G. sinensis. These results can provide a reference for growers in selecting Gleditsia varieties and provide insights into the industrial applications of Gleditsia endosperm products.

Analysis of differential mRNA and miRNA expression induced by heterogeneous grafting in Gleditsia sinensis.

Gleditsia sinensis Lam. is a multifaceted plant with medicinal, edible, chemical, timber, and ornamental applications. However, the effect of rootstocks on scions after grafting is still unclear. This study examined the mRNA and miRNA transcriptome among homografts, heterografts, and seedlings. GO enrichment analysis between seedlings and homograft/heterograft combinations revealed that biosynthesis, degradation, and transport were enriched. The KEGG enrichment results showed that plant hormone signal transduction and the plant MAPK signaling pathway were enriched in both seedlings and heterograft combinations. Through weighted correlation network analysis (WGCNA), the hub genes related to the content of plant hormones were obtained. Taking G. sinensis as the scion, there were 4594, 2887, 3429, and 5959 mRNAs that were specifically expressed in the grafted plants of G. sinensis/G. fera, G. sinensis/G. delavayi, G. sinensis/G. microphylla, and G. sinensis/G. japonica, respectively. The specifically expressed mRNA genes may participate in such processes and pathways as the rhythmic process, circadian rhythm, gibberellic-acid-mediated signaling pathway, and peptide-based amino acid modification. Additionally, 3, 16, 2, and 15 specifically expressed miRNAs were identified. This study examines the impact of grafting on gene expression in Gleditsia plants and establishes a foundation for the development of new resources and rootstock breeding.

Physiological responses to drought stress of three pine species and comparative transcriptome analysis of Pinus yunnanensis var. pygmaea.

Drought stress can significantly affect plant growth, development, and yield. Fewer comparative studies have been conducted between different species of pines, particularly involving Pinus yunnanensis var. pygmaea (P. pygmaea). In this study, the physiological indices, photosynthetic pigment and related antioxidant enzyme changes in needles from P. pygmaea, P. elliottii and P. massoniana under drought at 0, 7, 14, 21, 28 and 35 d, as well as 7 days after rehydration, were measured. The PacBio single-molecule real-time (SMRT) and Illumina RNA sequencing were used to uncover the gene expression differences in P. pygmaea under drought and rehydration conditions. The results showed that the total antioxidant capacity (TAOC) of P. pygmaea was significantly higher than P. massoniana and P. elliottii. TAOC showed a continuous increase trend across all species. Soluble sugar (SS), starch content and non-structural carbohydrate (NSC) of all three pines displayed a "W" pattern, declining initially, increasing, and then decreasing again. P. pygmaea exhibits stronger drought tolerance and greater recovery ability under prolonged drought conditions. Through the PacBio SMRT-seq, a total of 50,979 high-quality transcripts were generated, and 6,521 SSR and 5,561 long non-coding RNAs (LncRNAs) were identified. A total of 2310, 1849, 5271, 5947, 7710, and 6854 differentially expressed genes (DEGs) were identified compared to the control (Pp0D) in six pair-wise comparisons of treatment versus control. bHLH, NAC, ERF, MYB_related, C3H transcription factors (TFs) play an important role in drought tolerance of P. pygmaea. KEGG enrichment analysis and Gene set enrichment analysis (GSEA) analysis showed that P. pygmaea may respond to drought by enhancing metabolic processes such as ABA signaling pathway, alpha-linolenic acid. Weighted gene co-expression network analysis (WGCNA) revealed GST, CAT, LEC14B, SEC23 were associated with antioxidant enzyme activity and TAOC. This study provides a basis for further research on drought tolerance differences among coniferous species.

Characterization of the chloroplast genome of Gleditsia species and comparative analysis.

The genus Gleditsia has significant medicinal and economic value, but information about the chloroplast genomic characteristics of Gleditsia species has been limited. Using the Illumina sequencing, we assembled and annotated the whole chloroplast genomes of seven Gleditsia species (Gleditsia sinensis, Gleditsia japonica var. delavayi (G. delavayi), G. fera, G. japonica, G. microphylla, Fructus Gleditsiae Abnormalis (Zhu Yá Zào), G. microphylla mutant). The assembled genomes revealed that Gleditsia species have a typical circular tetrad structure, with genome sizes ranging from 162,746 to 170,907 bp. Comparative genomic analysis showed that most (65.8-75.8%) of the abundant simple sequence repeats in Gleditsia and Gymnocladus species were located in the large single copy region. The Gleditsia chloroplast genome prefer T/A-ending codons and avoid C/G-ending codons, positive selection was acting on the rpoA, rpl20, atpB, ndhA and ycf4 genes, most of the chloroplast genes of Gleditsia species underwent purifying selection. Expansion and contraction of the inverted repeat (IR)/single copy (SC) region showed similar patterns within the Gleditsia genus. Polymorphism analysis revealed that coding regions were more conserved than non-coding regions, and the IR region was more conserved than the SC region. Mutational hotspots were mostly found in intergenic regions such as "rps16-trnQ", "trnT-trnL", "ndhG-ndhI", and "rpl32-trnL" in Gleditsia. Phylogenetic analysis showed that G. fera is most closely related to G. sinensis,G. japonica and G. delavayi are relatively closely related. Zhu Yá Zào can be considered a bud mutation of the G. sinensis. The albino phenotype of G. microphylla mutant is not caused by variations in the chloroplast genome, and that the occurrence of the albino phenotype may be due to mutations in chloroplast-related genes involved in splicing or localization functions. This study will help us enhance our exploration of the genetic evolution and geographical origins of the Gleditsia genus.

Full-length transcriptome characterization and comparative analysis of Gleditsia sinensis.

As an economically important tree, Gleditsia sinensis Lam. is widely planted. A lack of background genetic information on G. sinensis hinders molecular breeding. Based on PacBio single-molecule real-time (SMRT) sequencing and analysis of G. sinensis, a total of 95,183 non-redundant transcript sequences were obtained, of which 93,668 contained complete open reading frames (ORFs), 2,858 were long non-coding RNAs (LncRNAs) and 18,855 alternative splicing (AS) events were identified. Genes orthologous to different Gleditsia species pairs were identified, stress-related genes had been positively selected during the evolution. AGA, AGG, and CCA were identified as the universal optimal codon in the genus of Gleditsia. EIF5A was selected as a suitable fluorescent quantitative reference gene. 315 Cytochrome P450 monooxygenases (CYP450s) and 147 uridine diphosphate (UDP)-glycosyltransferases (UGTs) were recognized through the PacBio SMRT transcriptome. Randomized selection of GsIAA14 for cloning verified the reliability of the PacBio SMRT transcriptome assembly sequence. In conclusion, the research data lay the foundation for further analysis of the evolutionary mechanism and molecular breeding of Gleditsia.

Comparative transcriptome analysis of dioecious floral development in Trachycarpus fortunei using Illumina and PacBio SMRT sequencing.

BACKGROUND: Trachycarpus fortunei is a plant with significant economic and ornamental value. Both male and female flowers of T. fortunei originate as bisexual flowers, and selective abortion occurs during floral development. However, the regulatory mechanisms underlying this process remain unclear in T. fortunei. In this study, transcriptome sequencing with Illumina and Pacific BioSciences (PacBio) single-molecule real-time (SMRT) platforms were used to investigate gene expression differences between male and female T. fortunei plants. RESULTS: A total of 833,137 full-length non-chimeric (FLNC) reads were obtained, and 726,846 high-quality full-length transcripts were identified. A total of 159 genes were differentially expressed between male and female flowers at all development stages. Some of the differentially expressed genes (DEGs) showed male bias, including serine/threonine-protein kinase (STPK), THUMP1 homolog and other genes. Through single-nucleotide polymorphisms(SNPs) identification, 28 genes were considered as potential sex-associated SNPs. Time-Ordered Gene Co-expression Network (TO-GCN) analysis revealed that MADS2 and MADS26 may play important roles in the development of female and male flowers T. fortune plants, respectively. CONCLUSIONS: These findings provide a genetic basis for flower development and differentiation in T. fortunei, and improve our understanding of the mechanisms underlying sexual differentiation in T. fortunei.