MiRNAs (microRNA) are 18–24 nt endogenous noncoding RNAs that regulate gene expression at the post-transcriptional level, including tissue-specific, developmental timing and evolutionary conservation gene expression. This study used high-throughput sequencing technology for the first time in Larix olgensis, predicted 78 miRNAs, including 12,229,003 reads sRNA, screened differentially expressed miRNAs. Predicting target genes was helpful for understanding the miRNA regulation function and obtained 333 corresponding target genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional annotation were analysed, mostly including nucleic acid binding, plant hormone signal transduction, pantothenate and CoA biosynthesis, and cellulose synthase. This study will lay the foundation for clarifying the complex miRNA-mediated regulatory network for growth and development. In view of this, spatio-temporal expression of miR396, miR950, miR164, miR166 and miR160 were analysed in Larix olgensis during the growth stages of not lignified, beginning of lignification, and completely lignified in different tissues (root, stem, and leaf) by quantitative real-time PCR (qRT-PCR). There were differences in the expression of miRNAs in roots, stems and leaves in the same growth period. At 60 days, miR160, miR166 and miR396–2 exhibited the highest expression in leaves. At 120 days, most miRNAs in roots and stems decreased significantly. At 180 days, miRNAs were abundantly expressed in roots and stems. Meanwhile, analysis of the expression of miRNAs in leaves revealed that miR396–2 was reduced as time went on, whereas other miRNAs increased initially and then decreased. On the other hand, in the stems, miR166–1 was increase, whereas other miRNAs, especially miR160, miR164, miR396 and miR950–1, first decreased and then increased. Similarly, in the roots, miR950–2 first decreased and then increased, whereas other miRNAs exhibited a trend of continuous increase. The present investigation included rapid isolation and identification of miRNAs in Larix olgensis through construction of a sRNA library using Solexa and predicted 78 novel miRNAs, which showed differential expression levels in different tissues and stages. These results provided a theoretical basis for further revealing the genetic regulation mechanism of miRNA in the growth and development of conifers and the verification of function in target genes.

BackgroundDrought stress in trees limits their growth, survival, and productivity, and it negatively affects the afforestation survival rate. Molecular responses to drought stress have been extensively studied in broad-leaved species, but studies on coniferous species are limited.ResultsOur study focused on the molecular responses to drought stress in a coniferous species, Larix olgensis A. Henry. Drought stress was simulated in one-year-old seedlings using 25% polyethylene glycol 6000. The drought stress response in these seedlings was assessed by analyzing select biochemical parameters, along with gene expression and metabolite profiles. The soluble protein content, peroxidase activity, and malondialdehyde content of L. olgensis were significantly changed during drought stress. Quantitative gene expression analysis identified a total of 8172 differentially expressed genes in seedlings processed after 24 h, 48 h, and 96 h of drought stress treatment. Compared with the gene expression profile of the untreated control, the number of up-regulated genes was higher than that of down-regulated genes, indicating that L. olgensis mainly responded to drought stress through positive regulation. Metabolite analysis of the control and stress-treated samples showed that under drought stress, the increased abundance of linoleic acid was the highest among up-regulated metabolites, which also included some saccharides. A combined analysis of the transcriptome and metabolome revealed that genes dominating the differential expression profile were involved in glutathione metabolism, galactose metabolism, and starch and sucrose metabolism. Moreover, the relative abundance of specific metabolites of these pathways was also altered. Thus, our results indicated that L. olgensis prevented free radical-induced damage through glutathione metabolism and responded to drought through sugar accumulation.ConclusionsThe soluble protein content, peroxidase activity, and malondialdehyde content of L. olgensis were significantly changed during drought stress. A total of 8172 differentially expressed genes in seedlings processed after drought stress treatment. A combined analysis of the transcriptome and metabolome revealed that genes dominating the differential expression profile were involved in glutathione metabolism, galactose metabolism, and starch and sucrose metabolism. Our results indicated that L. olgensis prevented free radical-induced damage through glutathione metabolism and responded to drought through sugar accumulation.

Background Larch (Larix gmelinii (Rupr.) Kuzen.) is an important timber and ecological tree species in northern China. Excellent germplasm resources have been acquired through time-consuming traditional breeding. Molecular breeding offers a promising approach to shorten the breeding cycle and achieve genetic improvements more efficiently. MicroRNAs (miRNAs) are non-coding, single-stranded small RNAs that primarily affect plant growth and stress resistance, including drought stress. However, the study of miRNAs in larch under drought stress has not been well explored. Results In this study, to investigate the function of Lol-miR11467 under PEG osmotic stress in larch, embryogenic callus tissue of Larix kaempferi 3 × L. gmelinii 9 was employed as the experimental material, serving as the explants for this study. Lol-miR11467was transferred into the explants using an Agrobacterium-mediated method to determine the physiological changes and survey gene expression changes in overexpressing Lol-miR11467 cell lines. The results showed that the fresh weight, peroxidase (POD), soluble protein and soluble sugar content of the overexpressing Lol-miR11467 were lower than that of the wild-type, while malondialdehyde (MDA) content increased under PEG osmoticstress. Transcriptome analysis showed that genes associated with phenylpropanoid metabolism, transcription factors, oxidoreductase, plant hormone signal transduction, glucose metabolism and bioprotective macromolecules were mainly downregulated in Lol-miR11467 cell lines. Conclusions Overall, these results indicated that the drought resistance of the overexpressing Lol-miR11467 cell lines was reduced. This study'sfindings might provide a foundation for understanding the molecular mechanisms of miRNAs under PEG osmotic stress in larch, potentially contributing to the development of strategies for improving plant resilience to environmental stresses.