Abstract
BackgroundThe Pythium splendens is a potentially useful organism for the synthesis of large amounts of eicosapentaenoic acid. Peak biomass and lipid accumulation do not occur at the same time and growth temperature has an effect on the fatty acid composition. Little is known about the pathway or the genes involved in growth, lipid synthesis or temperature resistance in P. splendens. Analysis of the transcriptome and expression profile data for P.splendensRBB-5 were used to extend genetic information for this strain and to contribute to a comprehensive understanding of the molecular mechanisms involved in specific biological processes.Methodology/Principal FindingsThis study used transcriptome assembly and gene expression analysis with short-read sequencing technology combined with a tag-based digital gene expression (DGE) system. Assembled sequences were annotated with gene descriptions, such as gene ontology (GO), clusters of orthologous group (COG) terms and KEGG orthology (KO) to generate 23,796 unigenes. In addition, we obtained a larger number of genes at different stages of fermentation (48, 100 and 148 h). The genes related to growth characteristics and lipid biosynthesis were analyzed in detail. Some genes associated with lipid and fatty acid biosynthesis were selected to confirm the digital gene expression (DGE) results by quantitative real-time PCR (qRT-PCR).Conclusion/SignificanceThe transcriptome improves our genetic understanding of P.splendensRBB-5 greatly and makes a large number of gene sequences available for further study. Notably, the transcriptome and DGE profiling data of P.splendensRBB-5 provide a comprehensive insight into gene expression profiles at different stages of fermentation and lay the foundation for the study of optimizing lipid content and growth speed at the molecular level.
Highlights
Eicosapentaenoic acid (EPA; 5,8,11,14,17-cis-eicosapentaenoic acid) is an v-3 C20-polyunsaturated fatty acid (PUFA) with a wide range of functions in biological systems
A study of the growth and biosynthesis of EPA by P. irregular [18] shows accumulation of v-3 PUFAs is greater in cultures grown at low temperature (12,14uC), which was verified by Mortierella [19]
We found that glycerol-3-phosphate dehydrogenase (GPDH) is expressed abundantly only in the 100 h library andglyceraldehyde-3-phosphate dehydrogenase (GAPDH) is expressed abundantly in the 100 h and 148 h libraries, which suggests growth and lipid biosynthesis are related to GAPDH and GPDH directly
Summary
Eicosapentaenoic acid (EPA; 5,8,11,14,17-cis-eicosapentaenoic acid) is an v-3 C20-polyunsaturated fatty acid (PUFA) with a wide range of functions in biological systems. The potential of a transgenic plant [11,12] for EPA production was studied owing to the high oil content in the seed, and several genes (D5, D6 and v-3 fatty acid desaturases etc.) were introduced into the plant, the yields of PUFAs are high but the growth period of the plant is lengthy and is constrained by geography. Some Pythium lipid synthesis genes are cloned and their functions are known (e.g. D5, D6 and D17 fatty acid desaturases) [21,22,23], but a comprehensive analysis of the mechanisms underlying growth and biosynthesis of lipids and EPA is needed to aid improvement of the EPA yield. Analysis of the transcriptome and expression profile data for P.splendensRBB-5 were used to extend genetic information for this strain and to contribute to a comprehensive understanding of the molecular mechanisms involved in specific biological processes
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