De Novo Sequencing and Transcriptome Analysis Reveal Genes' Specific Expression in Chinese Fir (Cunninghamia lanceolata) Callus

While the progress made in vitro culture of Chinese fir has produced satisfactory results, further improvements are warranted. To understand the mechanism of somatic embryogenesis (SE) in Chinese fir, we conducted phenotypic observations, physiological and biochemical measurements, and transcriptome analysis of embryonic (EC) and non-embryogenic callus (NEC) to provide a scientific basis for SE in this species. We found that EC and NEC showed significant morphological and physiological-biochemical indicators differences. Compared with NEC, EC had higher levels of soluble protein and proline and lower levels of malondialdehyde (MDA), peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT). Callus transcriptome sequencing assembled 152,229 unigenes, and 438 differentially expressed genes (DEGs) were screened, including transcription factor-related (TFs), DNA methylation-related, cell wall component protein, signal transduction-related, and stress response-related. GO and KEGG enrichment analyses of DEGs identified starch and sucrose, glutathione, and cysteine and methionine metabolism as the most representative pathways significantly enriched in EC and NEC genes and were associated with cell proliferation and embryogenesis. For the first time, the specific patterns of gene expression in Chinese fir callus were found through transcriptome comparison between callus, 16-year-old Chinese fir cambium, and drought-stressed tissue culture seedlings. In Chinese fir callus, 75.1% of genes were co-expressed in 16-year-old Chinese fir cambium and drought-stressed tissue culture seedlings, and 24.9% were only specifically expressed in callus. DEGs from EC and NEC indicated that 68.2 and 31.8% were co-expressed and specifically expressed, respectively. These results provided a basis for Chinese fir rapid propagation, which is expected to have theoretical and practical significance.