iTRAQ-based quantitative proteomics analysis of an immature high-oleic acid near-isogenic line of rapeseed

摘要

High-oleic acid rapeseed is a new and promising rapeseed variety because it produces oil with high-quality health and nutrient properties. To identify differentially expressed proteins in a high-oleic acid rapeseed line, self-bred seeds (20 to 35 days after pollination) of a high-and a low-oleic acid rapeseed near-isogenic line (oleic acid contents of 81.4 and 56.2%, respectively) were used as raw materials for iTRAQ (isobaric tags for relative and absolute quantitation) analysis. In total, 24,755 peptides, 14,455 unique peptides, and 4726 proteins were identified. Among them, 137 differentially expressed proteins were assigned pathway annotations, accounting for 3.84% of the total proteins (3568). Three differentially expressed proteins, PSaB (gi vertical bar 383930478), Lhcb6 (gi vertical bar 50313237), and ALD (gi vertical bar 15227981) were subjected to western blotting analyses. The results were consistent with the iTRAQ data, demonstrating the satisfactory quality of the iTRAQ experimental procedures and data. The transcript levels of 19 genes involved in fatty acid metabolism and photosynthesis were analyzed by qPCR. The transcript levels were consistent with the protein levels for 10 of these genes, but inconsistent for the other nine genes. Among the differentially expressed proteins were key enzymes in fatty acid biosynthesis and polyunsaturated fatty acid biosynthesis (acyl-CoA thioesterase, 3-ketoacyl-CoA reductase, omega 6 reductase, and acyl-CoA dehydrogenase), oleic acid biosynthesis (acyl-CoA thioesterase and omega 6 reductase), and seed lipid content (long-chain acyl-CoA synthetase). These results provide fundamental functional proteomics information for further breeding work on high-oleic acid rapeseed. This information will also be useful for research on the molecular mechanism of oleic acid synthesis.