Expression profiles of genes involved in fatty acid and lipid biosynthesis in developing seeds of Paeonia ostii.

BACKGROUND: Paeonia ostii seeds were identified as novel sources of edible plant oil with a high proportion of α-linolenic acid, a type of n-3 fatty acid with many health benefits. Due to the unreliability of seed oil content and quality, it is necessary to discover the mechanism underlying lipid biosynthesis in Paeonia ostii seeds. OBJECTIVES: This study aimed to identify the key genes involved in lipid biosynthesis in Paeonia ostii seeds by analyzing the relationship among the seed characteristics and the expression patterns of lipid genes in Paeonia ostii during seed development. METHODS: Preliminary research on Paeonia ostii seed development was carried out from 10 days after pollination until maturity, focusing on phenology, oil content and lipid profiles. In addition, we investigated the spatiotemporal expression of 36 lipid biosynthetic genes in Paeonia ostii by using quantitative real-time PCR. RESULTS: The results suggested that the development of Paeonia ostii seeds from pollination to maturity could be divided into three periods. The 36 lipid genes showed various spatiotemporal expression patterns and five gene groups with distinct temporal patterns during seed development were identified by clustering analysis of expression data. Furthermore, the relationships between gene expression and lipid/fatty acid accumulation and some candidate key lipid genes were discussed. CONCLUSIONS: This study provided the global patterns of fatty acid and lipid biosynthesis-related gene expression, which are critical to understanding the molecular basis of lipid biosynthesis and identifying the lipid accumulation rate-limiting genes during seed development.

Selection of Reliable Reference Genes for Gene Expression Studies of a Promising Oilseed Crop, Plukenetia volubilis, by Real-Time Quantitative PCR.

Real-time quantitative PCR (RT-qPCR) is a reliable and widely used method for gene expression analysis. The accuracy of the determination of a target gene expression level by RT-qPCR demands the use of appropriate reference genes to normalize the mRNA levels among different samples. However, suitable reference genes for RT-qPCR have not been identified in Sacha inchi (Plukenetia volubilis), a promising oilseed crop known for its polyunsaturated fatty acid (PUFA)-rich seeds. In this study, using RT-qPCR, twelve candidate reference genes were examined in seedlings and adult plants, during flower and seed development and for the entire growth cycle of Sacha inchi. Four statistical algorithms (delta cycle threshold (ΔCt), BestKeeper, geNorm, and NormFinder) were used to assess the expression stabilities of the candidate genes. The results showed that ubiquitin-conjugating enzyme (UCE), actin (ACT) and phospholipase A22 (PLA) were the most stable genes in Sacha inchi seedlings. For roots, stems, leaves, flowers, and seeds from adult plants, 30S ribosomal protein S13 (RPS13), cyclophilin (CYC) and elongation factor-1alpha (EF1α) were recommended as reference genes for RT-qPCR. During the development of reproductive organs, PLA, ACT and UCE were the optimal reference genes for flower development, whereas UCE, RPS13 and RNA polymerase II subunit (RPII) were optimal for seed development. Considering the entire growth cycle of Sacha inchi, UCE, ACT and EF1α were sufficient for the purpose of normalization. Our results provide useful guidelines for the selection of reliable reference genes for the normalization of RT-qPCR data for seedlings and adult plants, for reproductive organs, and for the entire growth cycle of Sacha inchi.