Identification of candidate genes regulating seed oil content by QTL mapping and transcriptome sequencing in Brassica napus.

Increasing oil production is a major goal in rapeseed (Brassica napus) molecular breeding programs. Identifying seed oil content (SOC)-related candidate genes is an important step towards achieving this goal. We performed quantitative trait locus (QTL) mapping of SOC in B. napus using a high-density SNP genetic map constructed from recombinant inbred lines and the Illumina InfiniumTM 60K SNP array. A total of 26 QTLs were detected in three years on A01, A03, A05, A06, A09, C01, C03 and C05, which accounted for 3.69%~18.47% of the phenotypic variation in SOC. Of these, 13 QTLs are reported here for the first time. 1713 candidate genes in the 26 QTLs confidence interval were obtained. We then identified differentially expressed genes (DEGs) between the high- and low-SOC accessions, to narrow down our focus to 21 candidate genes (Y1-Y21) related to SOC, and we will focus on 11 (Y1-Y11) candidate genes that contribute to the formation of high-SOC. In addition to providing insight into the genetic basis of SOC in B. napus, the loci identified and candidate genes in this study can be used in molecular breeding strategies to increase SOC in this important seed crop.

The Brassica napus fatty acid exporter FAX1-1 contributes to biological yield, seed oil content, and oil quality.

BACKGROUND: In the oilseed crop Brassica napus (rapeseed), various metabolic processes influence seed oil content, oil quality, and biological yield. However, the role of plastid membrane proteins in these traits has not been explored. RESULTS: Our genome-wide association study (GWAS) of 520 B. napus accessions identified the chloroplast membrane protein-localized FATTY ACID EXPORTER 1-1 (FAX1-1) as a candidate associated with biological yield. Seed transcript levels of BnaFAX1-1 were higher in a cultivar with high seed oil content relative to a low-oil cultivar. BnaFAX1-1 was localized to the plastid envelope. When expressed in Arabidopsis thaliana, BnaFAX1-1 enhanced biological yield (total plant dry matter), seed yield and seed oil content per plant. Likewise, in the field, B. napus BnaFAX1-1 overexpression lines (BnaFAX1-1-OE) displayed significantly enhanced biological yield, seed yield, and seed oil content compared with the wild type. BnaFAX1-1 overexpression also up-regulated gibberellic acid 4 (GA4) biosynthesis, which may contribute to biological yield improvement. Furthermore, oleic acid (C18:1) significantly increased in BnaFAX1-1 overexpression seeds. CONCLUSION: Our results indicated that the putative fatty acid exporter BnaFAX1-1 may simultaneously improve seed oil content, oil quality and biological yield in B. napus, providing new approaches for future molecular breeding.

Genome-Wide Identification and Expression Profiling of Monosaccharide Transporter Genes Associated with High Harvest Index Values in Rapeseed (Brassica napus L.).

Sugars are important throughout a plant's lifecycle. Monosaccharide transporters (MST) are essential sugar transporters that have been identified in many plants, but little is known about the evolution or functions of MST genes in rapeseed (Brassica napus). In this study, we identified 175 MST genes in B. napus, 87 in Brassica oleracea, and 83 in Brassica rapa. These genes were separated into the sugar transport protein (STP), polyol transporter (PLT), vacuolar glucose transporter (VGT), tonoplast monosaccharide transporter (TMT), inositol transporter (INT), plastidic glucose transporter (pGlcT), and ERD6-like subfamilies, respectively. Phylogenetic and syntenic analysis indicated that gene redundancy and gene elimination have commonly occurred in Brassica species during polyploidization. Changes in exon-intron structures during evolution likely resulted in the differences in coding regions, expression patterns, and functions seen among BnMST genes. In total, 31 differentially expressed genes (DEGs) were identified through RNA-seq among materials with high and low harvest index (HI) values, which were divided into two categories based on the qRT-PCR results, expressed more highly in source or sink organs. We finally identified four genes, including BnSTP5, BnSTP13, BnPLT5, and BnERD6-like14, which might be involved in monosaccharide uptake or unloading and further affect the HI of rapeseed. These findings provide fundamental information about MST genes in Brassica and reveal the importance of BnMST genes to high HI in B. napus.

Identification of candidate genes controlling oil content by combination of genome-wide association and transcriptome analysis in the oilseed crop Brassica napus.

BACKGROUND: Increasing seed oil content is one of the most important targets for rapeseed (Brassica napus) breeding. However, genetic mechanisms of mature seed oil content in Brassica napus (B. napus) remain little known. To identify oil content-related genes, a genome-wide association study (GWAS) was performed using 588 accessions. RESULTS: High-throughput genome resequencing resulted in 385,692 high-quality single nucleotide polymorphism (SNPs) with a minor allele frequency (MAF) > 0.05. We identified 17 loci that were significantly associated with seed oil content, among which 12 SNPs were distributed on the A3 (11 loci) and A1 (one loci) chromosomes, and five novel significant SNPs on the C5 (one loci) and C7 (four loci) chromosomes, respectively. Subsequently, we characterized differentially expressed genes (DEGs) between the seeds and silique pericarps on main florescences and primary branches of extremely high- and low-oil content accessions (HO and LO). A total of 64 lipid metabolism-related DEGs were identified, 14 of which are involved in triacylglycerols (TAGs) biosynthesis and assembly. Additionally, we analyzed differences in transcription levels of key genes involved in de novo fatty acid biosynthesis in the plastid, TAGs assembly and lipid droplet packaging in the endoplasmic reticulum (ER) between high- and low-oil content B. napus accessions. CONCLUSIONS: The combination of GWAS and transcriptome analyses revealed seven candidate genes located within the confidence intervals of significant SNPs. Current findings provide valuable information for facilitating marker-based breeding for higher seed oil content in B. napus.

Genome-Wide Identification and Comparative Expression Profile Analysis of the Long-Chain Acyl-CoA synthetase (LACS) Gene Family in Two Different Oil Content Cultivars of Brassica napus.

Long-chain acyl-CoA synthetase (LACS) is one of the key enzymes involved in fatty acid metabolism, including phospholipid biosynthesis, triacylglycerol (TAG) biosynthesis, and fatty acid ß-oxidation in plants. However, the characterization of LACSs family in seed oil biosynthesis of Brassica napus (B. napus) remains unknown. In the present study, we performed a comprehensive genome-wide analysis of this gene family in B. napus, and 34 B. napus LACS genes (BnaLACSs) were identified. Phylogenetic analysis classified the BnaLACS proteins into four groups (A, B, C, and D), which were supported by highly conserved gene structures and consensus motifs. RNA-Sequencing (RNA-Seq) and qRT-PCR combined analysis revealed that 18 BnaLACSs (BnaLACS1-2, 1-3, 1-4, 1-9, 1-10, 2-1, 2-2, 4-1, 4-2, 6-1, 6-2, 6-4, 7-1, 7-2, 8-1, 8-2, 9-3, and 9-4) were highly expressed in developmental seeds. Comparative expression analysis between extremely high oil content (P1-HO) and low oil content (P2-LO) B. napus cultivars revealed that BnaLACS6-4, BnaLACS9-3, and BnaLACS9-4 may be involved in fatty acid synthesis in chloroplast, and BnaLACS1-10 and 4-1 may play a vital role in lipid biosynthesis in B. napus, which is important for further seed oil accumulation in oilseed rape. The present study provides important information for functional characterization of BnaLACSs in seed oil metabolism in B. napus.

Characterization of Fatty Acid Exporters involved in fatty acid transport for oil accumulation in the green alga Chlamydomonas reinhardtii.

BACKGROUND: In the past few decades, microalgae biofuel has become one of the most interesting sources of renewable energy. However, the higher cost of microalgae biofuel compared to that of petroleum prevented microalgae biofuel production. Therefore, the research on increasing lipid productivity from microalgae becomes more important. The lipid production source, triacylglycerol biosynthesis in microalgae requires short chain fatty acids as substrates, which are synthesized in chloroplasts. However, the transport mechanism of fatty acids from microalgae chloroplasts to cytosol remains unknown. RESULTS: cDNAs from two homologs of the Arabidopsis fatty acid exporter 1 (FAX1) were cloned from Chlamydomonas reinhardtii and were named crfax1 and crfax2. Both CrFAXs were involved in fatty acid transport, and their substrates were mainly C16 and C18 fatty acids. Overexpression of both CrFAXs increased the accumulation of the total lipid content in algae cells, and the fatty acid compositions were changed under normal TAP or nitrogen deprivation conditions. Overexpression of both CrFAXs also increased the chlorophyll content. The MGDG content was decreased but the TAG, DAG, DGDG and other lipid contents were increased in CrFAXs overexpression strains. CONCLUSION: These results reveal that CrFAX1 and CrFAX2 were involved in mediating fatty acid export for lipids biosynthesis in C. reinhardtii. In addition, overexpression of both CrFAXs obviously increased the intracellular lipid content, especially the triacylglycerol content in microalgae, which provides a potential technology for the production of more biofuels using microalgae.