A systematic genome-wide analysis and screening of the NAC family genes related to wood formation in Cinnamomum camphora.

Many processes, such as growth, aging, and adaptation to abiotic stress, are regulated in plants by NAC transcription factors. In woody plants, NAC transcription factors acts as a primary switch that regulates secondary xylem development by activating various downstream transcription factors and modulating expression levels of genes involved in the synthesis of the secondary cell wall. Our team had previously sequenced the whole genome of the camphor tree (Cinnamomum camphora). Here, we performed a detailed analysis of the NAC gene family of C. camphora and examined its evolutionary history. The genomic sequences of 121 NAC genes of C. camphora were identified and classified into 20 subfamilies in 2 major classes based on the phylogenetic analysis and structural features. Expansion of the CcNAC gene family occurred mainly by fragment replication and was influenced by the purifying selection. By analyzing predicted interactions of the homologous AtNAC proteins, we identified five CcNACs that potentially regulate xylem development in C. camphora. RNA sequencing revealed distinct expression profiles of CcNACs in seven different plant tissues. Subcellular localization prediction revealed that 120, 3, and 2 CcNACs have biological functions in the nucleus, cytoplasm, and chloroplast, respectively. Furthermore, we verified expression patterns of five CcNACs (CcNAC012, CcNAC028, CcNAC055, CcNAC080, and CcNAC119) in various tissue types using qRT-PCR. Our results will facilitate further in-depth studies of the molecular mechanisms by which CcNAC transcription factors regulate wood formation and other processes in C. camphora.

Genome-Scale Identification, Classification, and Expression Profiling of MYB Transcription Factor Genes in Cinnamomum camphora.

The camphor tree (Cinnamomum camphora (L.) Presl.) is the representative species of subtropical evergreen broadleaved forests in eastern Asia and an important raw material for essential oil production worldwide. Although MYBs have been comprehensively characterized and their functions have been partially resolved in many plants, it has not been explored in C. camphora. In this study, 121 CcMYBs were identified on 12 chromosomes in the whole genome of C. camphora and found that CcMYBs were mainly expanded by segmental duplication. They were divided into 28 subgroups based on phylogenetic analysis and gene structural characteristics. In the promoter regions, numerous cis-acting elements were related to biological processes. Analysis of RNA sequencing data from seven tissues showed that CcMYBs exhibited different expression profiles, suggesting that they have various roles in camphor tree development. In addition, combined with the correlation analysis of structural genes in the flavonoid synthesis pathway, we identified CcMYBs from three subgroups that might be related to the flavonoid biosynthesis pathway. This study systematically analyzed CcMYBs in C. camphora, which will set the stage for subsequent research on the functions of CcMYBs during their lifetime and provide valuable insights for the genetic improvement of camphor trees.

Protoplast isolation and transient transformation system for Ginkgo biloba L.

Ginkgo biloba L. has a unique evolutionary status. Owing to its high medicinal and ornamental value, ginkgo has also recently become a research hotspot. However, the large genome and long juvenile period, as well as the lack of an effective genetic transformation system, have hindered gaining a full understanding of the comprehensive functions of ginkgo genes. At present, heterologous expression of genes in model plants is the primary method used in ginkgo-related research; however, these distant plant model relatives limit reliable interpretation of the results for direct applications in ginkgo breeding. To overcome these limitations, in this study, an efficient isolation and transient expression system for ginkgo protoplasts was established. A large number of intact and homogeneous ginkgo mesophyll protoplasts were isolated using 2% cellulase and 0.25% pectinase in 0.4 M mannitol. The activity of these protoplasts remained above 90% even after 24 h. Furthermore, when the concentration of the polyethylene glycol 4000 solution was 30%-40% (w/v), the transformation efficiency of the protoplasts reached 40%. Finally, the reliability of the system was verified using subcellular localization, transient overexpression, and protein interaction experiments with ginkgo genes, thereby providing a technical platform for the identification and analysis of ginkgo gene functions. The proposed method partially compensates for the limitations associated with the lack of a genetic transformation system and provides technical support to expand research on elucidating the functions of ginkgo genes.