Global warming increases sea temperature and leads to high temperature stress, which affects the yield and quality of Pyropia haitanensis. To understand the molecular mechanisms underlying high temperature stress in a high temperature tolerance strain Z-61, the iTRAQ technique was employed to reveal the global proteomic response of Z-61 under different durations of high temperature stress. We identified 151 differentially expressed proteins and classified them into 11 functional categories. The 4 major categories of these are protein synthesis and degradation, photosynthesis, defense response, and energy and carbohydrate metabolism. These findings indicated that photosynthesis, protein synthesis, and secondary metabolism are inhibited by heat to limit damage to a repairable level. As time progresses, misfolded proteins and ROS accumulate and lead to the up-regulation of molecular chaperones, proteases, and antioxidant systems. Furthermore, to cope with cells injured by heat, PCD works to remove them. Additionally, sulfur assimilation and cytoskeletons play essential roles in maintaining cellular and redox homeostasis. These processes are based on signal transduction in the phosphoinositide pathway and multiple ways to supply energy. Conclusively, Z-61 establishes a new steady-state balance of metabolic processes and survives under higher temperature stress.
Rehmannia glutinosa is a common bulk medicinal material that has been widely used in China due to its active ingredients. Acteoside, one of the ingredients, has antioxidant, antinephritic, anti-inflammatory, hepatoprotective, immunomodulatory, and neuroprotective effects, is usually selected as a quality-control component for R. glutinosa herb in the Chinese Pharmacopeia. The acteoside biosynthesis pathway in R. glutinosa has not yet been clearly established. Herein, we describe the establishment of a genetic transformation system for R. glutinosa mediated by Agrobacterium rhizogenes. We screened the optimal elicitors that markedly increased acteoside accumulation in R. glutinosa hairy roots. We found that acteoside accumulation dramatically increased with the addition of salicylic acid (SA); the optimal SA dose was 25 μmol/L for hairy roots. RNA-seq was applied to analyze the transcriptomic changes in hairy roots treated with SA for 24 h in comparison with an untreated control. A total of 3,716, 4,018, and 2,715 differentially expressed transcripts (DETs) were identified in 0 h-vs.-12 h, 0 h-vs.-24 h, and 12 h-vs.-24 h libraries, respectively. KEGG pathway-based analysis revealed that 127 DETs were enriched in “phenylpropanoid biosynthesis.” Of 219 putative unigenes involved in acteoside biosynthesis, 54 were found to be up-regulated at at least one of the time points after SA treatment. Selected candidate genes were analyzed by quantitative real-time PCR (qRT-PCR) in hairy roots with SA, methyl jasmonate (MeJA), AgNO3 (Ag+), and putrescine (Put) treatment. All genes investigated were up-regulated by SA treatment, and most candidate genes were weakly increased by MeJA to some degree. Furthermore, transcription abundance of eight candidate genes in tuberous roots of the high-acteoside-content (HA) cultivar QH were higher than those of the low-acteoside-content (LA) cultivar Wen 85-5. These results will pave the way for understanding the molecular basis of acteoside biosynthesis in R. glutinosa, and can serve as a basis for future validation studies.
BackgroundPyropia haitanensis is an economically important mariculture crop in China and is also valuable in life science research. However, the lack of genetic information of this organism hinders the understanding of the molecular mechanisms of specific traits. Thus, high-throughput sequencing is needed to generate a number of transcriptome sequences to be used for gene discovery and molecular marker development.ResultsIn this study, high-throughput sequencing was used to analyze the global transcriptome of P. haitanensis. Approximately 103 million 90 bp paired-end reads were generated using an Illumina HiSeq 2000. De novo assembly with paired-end information yielded 24,575 unigenes with an average length of 645 bp. Based on sequence similarity searches with known proteins, a total of 16,377 (66.64%) genes were identified. Of these annotated unigenes, 5,471 and 9,168 unigenes were assigned to gene ontology and clusters of orthologous groups, respectively. Searching against the KEGG database indicated that 12,167 (49.51%) unigenes mapped to 124 KEGG pathways. Among the carbon fixation pathways, almost all the essential genes related to the C3- and C4-pathways for P. haitanensis were discovered. Significantly different expression levels of three key genes (Rubisco, PEPC and PEPCK) in different lifecycle stages of P. haitanensis indicated that the carbon fixation pathway in the conchocelis and thallus were different, and the C4-like pathway might play important roles in the conchocelis stage. In addition, 2,727 cSSRs loci were identified in the unigenes. Among them, trinucleotide SSRs were the dominant repeat motif (87.17%, 2,377) and GCC/CCG motifs were the most common repeats (60.07%, 1,638). High quality primers to 824 loci were designed and 100 primer pairs were randomly evaluated in six strains of P. haitanensis. Eighty-seven primer pairs successfully yielded amplicons.ConclusionThis study generated a large number of putative P. haitanensis transcript sequences, which can be used for novel gene discovery and gene expression profiling analyses under different physiological conditions. A number of the cSSR markers identified can be used for molecular markers and will facilitate marker assisted selection in P. haitanensis breeding. These sequences and markers will provide valuable resources for further P. haitanensis studies.
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