Genome sequencing is widely recognized as a fundamental pillar in genetic research and legal studies of biological phenomena, providing essential insights for genetic investigations and legal analyses of biological events. The field of genome sequencing has experienced significant progress due to rapid improvements in scientific and technological developments. These advancements encompass not only significant improvements in the speed and quality of sequencing but also provide an unparalleled opportunity to explore the subtle complexities of genomes, particularly in the context of rare species. Such a wide range of possibilities has successfully supported the validation of plant gene functions and the refinement of precision breeding methodologies. This expanded scope now includes a comprehensive exploration of the current state and conservation efforts of gymnosperm gene sequencing, offering invaluable insights into their genomic landscapes. This comprehensive review elucidates the trajectory of development and the diverse applications of genome sequencing. It encompasses various domains, including crop breeding, responses to abiotic stress, species evolutionary dynamics, biodiversity, and the unique challenges faced in the conservation and utilization of gymnosperms. It highlights both ongoing challenges and the unveiling of forthcoming developmental trajectories.

Cunninghamia lanceolata (C. lanceolata) is an important timber tree species in southern China that requires gene function studies to understand its traits. In this study, we investigated the callus induction rates of immature zygotic embryos from reciprocal hybrids between genotypes B46 and B49. With zygotic embryo development, the callus induction rates showed an increasing trend, followed by a decreasing trend. Moreover, the rate of callus induction in genotype B46 × B49 immature zygotic embryos was greater than in genotype B49 × B46. Callus from C. lanceolata with genotype B46 × B49 was selected as the donor material for protoplast isolation. By using an enzymatic digestion solution containing cellulase, macerozyme, and pectinase, combined with an osmotic stabilizer, we obtained 9.76 × 10⁶ protoplasts/mL with 92.7% viability. We subsequently transformed plasmids into C. lanceolata callus protoplasts and observed the location of the H2B-eYFP fusion protein in the nucleus. To achieve transient transfection of C. lanceolata callus protoplasts, we compared transfection efficiencies at different concentrations of PEG4000, PEG6000, or PEG8000 in a modified MMg solution. We found that 20% (w/v) PEG6000 mediated the transient transfection of C. lanceolata callus protoplasts with a 6.70% efficiency. This study provides a technical foundation for future research on transient transfection and functional analysis of C. lanceolata genes.

Hemlocks (Pinaceae: Tsuga) are widely distributed in North America and East Asia, forming a reticulate evolutionary structure in East Asia with significant ecological importance. To clarify the chloroplast genome characteristics and phylogenetic relationships among Tsuga species, we analyzed the chloroplast genomes of T. chinensis var. tchekiangensis, T. chinensis, T. diversifolia, T. dumosa, T. forrestii, and T. sieboldii, performing associated phylogenetic analyses. The results reveal that the chloroplast genome lengths among the six Tsuga species vary from 120,520 to 121,010 bp, encompassing about 108 to 112 genes, including 35/32 tRNA genes and 4 rRNA genes. A codon usage analysis highlighted a preference for A/U-ending codons, and all six nucleotide types have A/T bases and a prevalence of mononucleotides. Notably, all Tsuga species exhibit inverted repeat (IR) contractions and possess unique hexanucleotides absent in the other species of Pinaceae, potentially making them more susceptible to gene recombination or rearrangement during evolution. While most variations are observed in non-coding regions, particularly in intergenic fragments, substantial variation sites are also present within the genes. The phylogenetic tree, constructed using chloroplast genomes, substantiates the sister taxa relationship between Tsuga and Nothotsuga. Furthermore, it confirms that T. chinensis var. tchekiangensis exhibits a closer relationship with T. forrestii than with T. chinensis. These findings not only provide partial evidence that T. chinensis may not constitute a monophyletic species but also underscore the necessity of reevaluating the taxonomic status of T. chinensis var. tchekiangensis. In addition, while the RSCU cluster analysis is basically consistent with the phylogenetic analysis, it also highlights a distinct differentiation between Nothotsuga and Tsuga. This study not only provides molecular-level phylogenetic classification evidence of Pinaceous genera via chloroplast genome analyses but also offers compelling evidence for further exploring the relationships and species delimitation among the hemlocks of East Asia.

Sulfated peptides, such as PSK, PSY, CIF, and RGF, are crucial regulators of plant growth, development, and stress responses, with their activity dependent on post-translational tyrosine sulfation by tyrosylprotein sulfotransferase (TPST). This study explores the evolutionary history and the interaction mechanisms between TPST and sulfated peptides in plants. Systematic analyses of multi-species genomes show that TPST can be traced back to the chlorophyte lineage, whereas PSK, a sulfated peptide, appears to have emerged in gymnosperms. TPST is evolutionarily conserved, typically present in low copy numbers across plant lineages, while its peptide substrates have expanded in angiosperms. In Liriodendron chinense, TPST-sulfated peptide gene promoters are enriched with cis-regulatory elements linked to abscisic acid, gibberellin responsiveness, and anaerobic induction. Synteny analyses revealed collinearity between sulfated peptide genes in L. chinense, Magnolia biondii, Arabidopsis thaliana, and Populus trichocarpa, but not with Oryza sativa. Molecular docking identified key TPST-PSK interaction sites in the sulfotransferase domain, with several critical residues facilitating binding. Transcriptomic and co-expression network analyses revealed that LcTPST was expressed at lower levels than its peptide precursor genes, while LcPSK2 remained highly expressed after the torpedo stage of somatic embryogenesis. Stress conditions significantly increased PSK-associated module connectivity, enriched in transcription factors such as WRKY, bHLH, bZIP, and MADS. This study provides insights into the evolutionary, structural, and regulatory aspects of the TPST-sulfated peptide system in plants.