Plant-based oils are valuable agricultural products, and seed oil content (SOC) is the major yield component in oil crops. Increasing SOC has been successfully targeted through the selection and genetic modification of oil biosynthesis. The SOC in rapeseed declined during the seed maturation and eventually caused the final accumulated seed oil quantity. However, genes involved in oil degradation during seed maturity are not deeply studied so far. We performed a candidate gene association study using a worldwide collection of rapeseed germplasm. We identified SEED FATTY ACID REDUCER (SFAR) genes, which had a significant effect on SOC and fatty acid (FA) composition. SFAR genes belong to the GDSL lipases, and GDSL lipases have a broad range of functions in plants. After quantification of gene expression using RNA-seq and quantitative PCR, we used targeted (CRISPR-Cas mediated) and random (chemical) mutagenesis to modify turnover rates of seed oil in winter rapeseed. For the first time, we demonstrate significant increase of SOC in a crop after knocking out members of the BnSFAR4 and BnSFAR5 gene families without pleiotropic effects on seed germination, vigour and oil mobilization. Our results offer new perspectives for improving oil yield by targeted mutagenesis.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/pbi.13381

Elevating seed oil content in a polyploid crop by induced mutations in SEED FATTY ACID REDUCER genes.

Engineering efforts targeted at increasing ethanol by modifying the central fermentative metabolism of Clostridium thermocellum have been variably successful. Here, we aim to understand this variation by a multifaceted approach including genomic and transcriptomic analysis combined with chemostat cultivation and high solids cellulose fermentation. Three strain lineages comprising 16 strains total were examined. Two strain lineages in which genes involved in pathways leading to organic acids and/or sporulation had been knocked out resulted in four end-strains after adaptive laboratory evolution (ALE). A third strain lineage recapitulated mutations involving adhE that occurred spontaneously in some of the engineered strains. Contrary to lactate dehydrogenase, deleting phosphotransacetylase (pta, acetate) negatively affected steady-state biomass concentration and caused increased extracellular levels of free amino acids and pyruvate, while no increase in ethanol was detected. Adaptive laboratory evolution (ALE) improved growth and shifted elevated levels of amino acids and pyruvate towards ethanol, but not for all strain lineages. Three out of four end-strains produced ethanol at higher yield, and one did not. The occurrence of a mutation in the adhE gene, expanding its nicotinamide-cofactor compatibility, enabled two end-strains to produce more ethanol. A disruption in the hfsB hydrogenase is likely the reason why a third end-strain was able to make more ethanol. RNAseq analysis showed that the distribution of fermentation products was generally not regulated at the transcript level. At 120 g/L cellulose loadings, deletions of spo0A, ldh and pta and adaptive evolution did not negatively influence cellulose solubilization and utilization capabilities. Strains with a disruption in hfsB or a mutation in adhE produced more ethanol, isobutanol and 2,3-butanediol under these conditions and the highest isobutanol and ethanol titers reached were 5.1 and 29.9 g/L, respectively. Modifications in the organic acid fermentative pathways in Clostridium thermocellum caused an increase in extracellular pyruvate and free amino acids. Adaptive laboratory evolution led to improved growth, and an increase in ethanol yield and production due a mutation in adhE or a disruption in hfsB. Strains with deletions in ldh and pta pathways and subjected to ALE demonstrated undiminished cellulolytic capabilities when cultured on high cellulose loadings.

/pdf/metabolic-and-evolutionary-responses-of-clostridium-2hmk79t7en.pdf

Metabolic and evolutionary responses of Clostridium thermocellum to genetic interventions aimed at improving ethanol production

Terpenoids are a diverse group of over 55,000 compounds with potential applications as advanced fuels, bulk and fine chemicals, pharmaceutical ingredients, agricultural chemicals, etc. To facilitate their bio-based production, there is a need for plug-and-play hosts, capable of high-level production of different terpenoids. Here we engineer Yarrowia lipolytica platform strains for the overproduction of mono-, sesqui-, di-, tri-, and tetraterpenoids. The monoterpene platform strain was evaluated by expressing Perilla frutescens limonene synthase, which resulted in limonene titer of 35.9 mg/L and was 100-fold higher than when the same enzyme was expressed in the strain without mevalonate pathway improvement. Expression of Callitropsis nootkatensis valencene synthase in the sesquiterpene platform strain resulted in 113.9 mg/L valencene, an 8.4-fold increase over the control strain. Platform strains for production of squalene, complex triterpenes, or diterpenes and carotenoids were also constructed and resulted in the production of 402.4 mg/L squalene, 22 mg/L 2,3-oxidosqualene, or 164 mg/L β-carotene, respectively. The presented terpenoid platform strains can facilitate the evaluation of terpenoid biosynthetic pathways and are a convenient starting point for constructing efficient cell factories for the production of various terpenoids. The platform strains and exemplary terpenoid strains can be obtained from Euroscarf.

/pdf/yarrowia-lipolytica-strains-engineered-for-the-production-of-3bjv88wrbn.pdf

Yarrowia lipolytica Strains Engineered for the Production of Terpenoids.

A la suite d'un appel d'offres du ministere federal de la Recherche et de l'Education (BMBF), un projet d'amelioration de la valeur nutritionnelle du colza (Brassica napus) pour des debouches alimentaires ou de nutrition humaine a ete initie en automne 1999. Un projet integre allant de la recherche, la caracterisation, l'isolement et le transfert de genes d'interet en selection, dans des varietes a haut potentiel et ayant des caracteristiques agronomiques interessantes, est actuellement conduit par un groupe de partenaires (figure 2). Des etudes economiques, des essais, ainsi que des analyses nutritionnelles pour des qualites nouvelles sont conduites dans ce programme. Les aspects de qualite des semences en relation avec la couleur des teguments de la graine, la composition en huile, en lecithine ainsi que les fractions proteiques et les antioxydants, tels que les tocopherols et le resveratol, seront ameliores ches Brassica napus.

NAPUS 2000 rapeseed (Brassica napus) breeding for improved human nutrition : Lipids, fats, and oils: opportunities and responsabilities in the new century

Non-model bacteria like Pseudomonas putida, Lactococcus lactis and other species have unique and versatile metabolisms, offering unique opportunities for Synthetic Biology (SynBio). However, key genome editing and recombineering tools require optimization and large-scale multiplexing to unlock the full SynBio potential of these bacteria. In addition, the limited availability of a set of characterized, species-specific biological parts hampers the construction of reliable genetic circuitry. Mining of currently available, diverse bacteriophages could complete the SynBio toolbox, as they constitute an unexplored treasure trove for fully adapted metabolic modulators and orthogonally-functioning parts, driven by the longstanding co-evolution between phage and host.

/pdf/exploring-the-synthetic-biology-potential-of-bacteriophages-4hkii2a0dh.pdf

Exploring the synthetic biology potential of bacteriophages for engineering non-model bacteria

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. 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. 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.

/pdf/identification-of-candidate-genes-controlling-oil-content-by-186g3fp1o5.pdf

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

Oilseed rape (Brassica napus) is the second largest oilseed crop worldwide. As an architecture component of B. napus, thickness of pod canopy (TPC) plays an important role in yield formation, especially under high-density cultivation conditions. However, the mechanisms underlying the regulation of TPC remain unclear. RNA and microRNA (miRNA) profiling of two groups of B. napus lines with significantly different TPC at the bolting with a tiny bud stage revealed differential expressions of numerous genes involved in nitrogen-related pathways. Expression of several nitrogen-related response genes, including ASP5, ASP2, ASN3, ATCYSC1, PAL2, APT2, CRTISO, and COX15, was dramatically changed in the thick TPC lines compared to those in the thin TPC lines. Differentially expressed miRNAs also included many involved in nitrogen-related pathways. Expression of most target genes was negatively associated with corresponding miRNAs, such as miR159, miR6029, and miR827. In addition, 12 (including miR319, miR845, and miR158) differentially expressed miRNAs between two plant tissues sampled (stem apex and flower bud) were identified, implying that they might have roles in determining overall plant architecture. These results suggest that nitrogen signaling may play a pivotal role in regulating TPC in B. napus.

https://www.mdpi.com/2073-4425/10/8/591/pdf

Joint RNA-Seq and miRNA Profiling Analyses to Reveal Molecular Mechanisms in Regulating Thickness of Pod Canopy in Brassica napus

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.

/pdf/the-brassica-napus-fatty-acid-exporter-fax1-1-contributes-to-54895l6ngs.pdf

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

Study on potential microbial community to the waste water treatment from bauxite desilication process

In recent years, significant efforts have been made to study changes in the levels of air pollutants at regional and urban scales, and changes in bioaerosols during air pollution events have attracted increasing attention. In this study, the bacterial structure of PM2.5 was analysed under different environmental conditions during hazy and non-hazy periods in Guilin. A total of 32 PM2.5 samples were collected in December 2020 and July 2021, and the microbial community structures were analysed using high-throughput sequencing methods. The results show that air pollution and climate change alter the species distribution and community diversity of bacteria in PM2.5, particularly Sphingomonas and Pseudomonas. The structure of the bacterial community composition is related to diurnal variation, vertical height, and urban area and their interactions with various environmental factors. This is a comprehensive study that characterises the variability of bacteria associated with PM2.5 in a variety of environments, highlighting the impacts of environmental effects on the atmospheric microbial community. The results will contribute to our understanding of haze trends in China, particularly the relationship between bioaerosol communities and the urban environment. 

https://link.springer.com/content/pdf/10.1007/s10453-022-09777-0.pdf

Qiang Huo

Papers

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

Joint RNA-Seq and miRNA Profiling Analyses to Reveal Molecular Mechanisms in Regulating Thickness of Pod Canopy in Brassica napus

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

Study on potential microbial community to the waste water treatment from bauxite desilication process

Comparison of bacterial community structure in PM2.5 during hazy and non-hazy periods in Guilin, South China