Background: Pyropia 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. Results: In 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. Conclusion: This 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.

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.

Pyropia haitanensis, a high-yield commercial seaweed in China, is currently undergoing increasing levels of high-temperature stress due to gradual global warming. The mechanisms of plant responses to high temperature stress vary with not only plant type but also the degree and duration of high temperature. To understand the mechanism underlying thermal tolerance in P. haitanensis, gene expression and regulation in response to short- and long-term temperature stresses (SHS and LHS) was investigated by performing genome-wide high-throughput transcriptomic sequencing for a high temperature tolerant strain (HTT). A total of 14,164 differential expression genes were identified to be high temperature-responsive in at least one time point by high-temperature treatment, representing 41.10% of the total number of unigenes. The present data indicated a decrease in the photosynthetic and energy metabolic rates in HTT to reduce unnecessary energy consumption, which in turn facilitated in the rapid establishment of acclimatory homeostasis in its transcriptome during SHS. On the other hand, an increase in energy consumption and antioxidant substance activity was observed with LHS, which apparently facilitates in the development of resistance against severe oxidative stress. Meanwhile, ubiquitin-mediated proteolysis, brassinosteroids, and heat shock proteins also play a vital role in HTT. The effects of SHS and LHS on the mechanism of HTT to resist heat stress were relatively different. The findings may facilitate further studies on gene discovery and the molecular mechanisms underlying high-temperature tolerance in P. haitanensis, as well as allow improvement of breeding schemes for high temperature-tolerant macroalgae that can resist global warming.

Chinese coastal waters are subject to eutrophication because of rapid economic development and population growth. To study the effects of nutrient enrichment on the growth and quality of Pyropia haitanensis, the thalli of a red-brown (Z-61) strain and a green (Z-26) strain of P. haitanensis were cultured semi-continuously for 15 days in four media with different nitrogen and phosphorus concentrations. High-nutrient conditions had no significant effect on the growth rate of Z-26 and Z-61. Higher concentrations of nutrients increased the chlorophyll a content of Z-61 and Z-26 by 116.7 and 34.2%, respectively. High concentrations of nutrients also significantly increased the average phycoerythrin and phycocyanin contents by 57.4 and 56.0%, respectively (strain Z-61); and by 37.7 and 44.7%, respectively (strain Z-26). As the nitrogen and phosphorus concentrations increased, the mean contents of total protein, essential amino acids, and flavor amino acids increased by 130.0, 102.5, and 341%, respectively, for strain Z-61, and by 25.1, 23.9, and 497%, respectively, for strain Z-26. Under high-nutrient conditions, the mean total amino acids content was 40.7% higher in Z-26 than in Z-61; the total flavor amino acids content was 117% higher in Z-61 than in Z-26; but there was no significant difference in the total essential amino acids content between the two strains. Interestingly, under low-nutrient conditions, Z-26 had a higher nutritional value than Z-61, but Z-61 tasted better than Z-26. Based on the results of these laboratory experiments, nutrient enrichment might enhance the market value of P. haitanensis by significantly improving its food quality.