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Selecting low-nitrogen(N)-tolerant maize hybrids represent an effective approach to enhancing nitrogen use efficiency grain yield. However, the impact of nitrogen fertilization on protein accumulation in low-N-tolerant hybrids remain insufficiently explored. In this paper, a two-year field orientation trial was conducted at four nitrogen fertilizer rate with the different low-N-tolerant maize hybrids. The effect of nitrogen fertilization on the accumulation of protein and its fractions different kernels positions of different low-N-tolerant maize hybrids was studied. The results showed that the protein yield of ZH311 maize kernels was significantly higher than that of XY508, especially under low-N conditions (0N and 150N), and was 25.7%-36.2% higher than that of XY508. There was a significant correlation between protein yield and the accumulation of crude protein and protein fractions. Compared with XY508, the crude protein of ZH311 entered the rapid growth stage later and lasted for a relatively shorter period, but it was 50.8%-53.0% higher due to its higher accumulation rates (v2 and v3) in its middle and late stages, especially in the apical grains. Under low-N conditions, the difference in crude protein accumulation between the apical and basal-middle kernels of ZH311 was only 4.3-8.2%, whereas the difference in XY508 was 29.9-37.3%, suggesting that low-N-tolerant maize hybrids improve protein yield by increasing the accumulation of proteins and their fractions in the apical kernels. Nitrogen fertilization had a greater effect on protein accumulation and yield in XY508, especially on the top kernel and protein yield. In the future, more attention should be paid to the effect of apical kernels when breeding high-quality maize hybrids tolerant to low nitrogen.

Gibberellic acid (GA3) is a vital plant growth hormone widely used in agriculture. Currently, GA3 production relies on liquid fermentation by the filamentous fungus Fusarium fujikuroi. However, the lack of an effective selection marker recycling system hampers the application of metabolic engineering technology in F. fujikuroi, as multiple-gene editing and positive-strain screening still rely on a limited number of antibiotics. In this study, we developed a strategy using pyr4-blaster and CRISPR/Cas9 tools for recycling orotidine-5′-phosphate decarboxylase (Pyr4) selection markers. We demonstrated the effectiveness of this method for iterative gene integration and large gene-cluster deletion. We also successfully improved GA3 titers by overexpressing geranylgeranyl pyrophosphate synthase and truncated 3-hydroxy-3-methyl glutaryl coenzyme A reductase, which rewired the GA3 biosynthesis pathway. These results highlight the efficiency of our established system in recycling selection markers during iterative gene editing events. Moreover, the selection marker recycling system lays the foundation for further research on metabolic engineering for GA3 industrial production.

Using supplementary cementitious materials in concrete production makes it eco-friendly by decreasing cement usage and the corresponding CO2 emissions. One key measure of concrete’s durability performance is its porosity. An empirical prediction of the porosity of high-performance concrete with added cementitious elements is the goal of this work, which employs machine learning approaches. Binder, water/cement ratio, slag, aggregate content, superplasticizer (SP), fly ash, and curing conditions were considered as inputs in the database. The aim of this study is to create ML models that could evaluate concrete porosity. Gene expression programming (GEP) and multi-expression programming (MEP) were used to develop these models. Statistical tests, Taylor’s diagram, R 2 values, and the difference between experimental and predicted readings were the metrics used to evaluate the models. With R 2 = 0.971, mean absolute error (MAE) = 0.348%, root mean square error (RMSE) = 0.460%, and Nash–Sutcliffe efficiency (NSE) = 0.971, the MEP provided a slightly better-fitted model and improved prediction performance when contrasted with the GEP, which had R 2 = 0.925, MAE = 0.591%, RMSE = 0.745%, and NSE = 0.923. Binder, water/binder ratio, curing conditions, and aggregate content had a direct (positive) relationship with the porosity of concrete, while SP, fly ash, and slag had an indirect (negative) association, according to the SHapley Additive exPlanations study.

Abscisic acid (ABA) is an important plant growth regulator with broad applications in agriculture, forestry, and other fields. Currently, the industrial production of ABA primarily relies on microbial fermentation using Botrytis cinerea, but its genetic toolbox is limited. To address this, we first screened 10 strong constitutive promoters from the genome of B. cinerea through transcriptomic analysis. The expression levels of the promoters covered a range of 3–4 orders of magnitude according to the measured β-glucuronidase activity. Subsequently, four promoters of different strength were used to balance the cofactor supply in B. cinerea. Overexpression of NADH kinase using the medium-strength promoter Pef1a significantly enhanced ABA production, resulting in a 32.26 % increase compared to the control. Finally, by combining promoter engineering with a push-pull strategy, we optimized the biosynthesis of ABA. The recombinant strain Pthi4:hmgr-Pef1a:a4, overexpressing HMGR under the Pthi4 promoter and Bcaba4 under the Pef1a promoter, achieved an ABA titer of 1.18 g/L, a 58.92 % increase. To our best knowledge, this is the first constitutive promoter library suitable for B. cinerea, providing important tools for the industrial production of ABA.

Marble cement (MC) is a new binding material for concrete, and the strength assessment of the resulting materials is the subject of this investigation. MC was tested in combination with rice husk ash (RHA) and fly ash (FA) to uncover its full potential. Machine learning (ML) algorithms can help with the formulation of better MC-based concrete. ML models that could predict the compressive strength (CS) of MC-based concrete that contained FA and RHA were built. Gene expression programming (GEP) and multi-expression programming (MEP) were used to build these models. Additionally, models were evaluated by calculating R 2 values, carrying out statistical tests, creating Taylor’s diagram, and comparing theoretical and experimental readings. When comparing the MEP and GEP models, MEP yielded a slightly better-fitted model and better prediction performance (R 2 = 0.96, mean absolute error = 0.646, root mean square error = 0.900, and Nash–Sutcliffe efficiency = 0.960). According to the sensitivity analysis, the prediction of CS was most affected by curing age and MC content, then by FA and RHA contents. Incorporating waste materials such as marble powder, RHA, and FA into building materials can help reduce environmental impacts and encourage sustainable development.

Abstract Background α-Humulene is an important biologically active sesquiterpene, whose heterologous production in microorganisms is a promising alternative biotechnological process to plant extraction and chemical synthesis. In addition, the reduction of production expenses is also an extremely critical factor in the sustainable and industrial production of α-humulene. In order to meet the requirements of industrialization, finding renewable substitute feedstocks such as low cost or waste substrates for terpenoids production remains an area of active research. Results In this study, we investigated the feasibility of peroxisome-engineering strain to utilize waste cooking oil (WCO) for high production of α-humulene while reducing the cost. Subsequently, transcriptome analysis revealed differences in gene expression levels with different carbon sources. The results showed that single or combination regulations of target genes identified by transcriptome were effective to enhance the α-humulene titer. Finally, the engineered strain could produce 5.9 g/L α‐humulene in a 5‐L bioreactor. Conclusion To the best of our knowledge, this is the first report that converted WCO to α-humulene in peroxisome-engineering strain. These findings provide valuable insights into the high-level production of α-humulene in Y. lipolytica and its utilization in WCO bioconversion. Graphical Abstract

The reliance of the transport sector on fossil fuels has led to increasing concerns related to sustainability and environmental impact. Growing petroleum shortages and global climate change caused by greenhouse gas emissions have given an impetus to the development of renewable and low-carbon energy sources. Fatty acid-derived hydrocarbons are an attractive drop-in biofuel to substitute fossil fuels consumed by the transport sector, especially heavy vehicles. Compared to traditional biofuels such as ethanol and biodiesel, fatty acid-derived hydrocarbons have higher energy density and better compatibility with existing storage and transport systems. As a common microbial platform for biofuel production, yeasts have been engineered to produce fatty acid-derived hydrocarbons. Here, we attempted to comprehensively review the latest advances in metabolic engineering strategies for the production of fatty acid-derived hydrocarbons in the yeasts Saccharomyces cerevisiae and Yarrowia lipolytica. The heterologous hydrocarbon synthesis pathways and combinational metabolic engineering strategies have been summarized, followed by a discussion of future research directions for the development of yeasts as industrial hydrocarbon producers.

Due to their vital physiological functions, nutritional fatty acids have great potential as nutraceutical food supplements for preventing an array of diseases such as inflammation, depression, arthritis, osteoporosis, diabetes and cancer. Microbial biosynthesis of fatty acids follows the trend of sustainable development, as it enables green, environmentally friendly and efficient production. As a natural oleaginous yeast, Yarrowia lipolytica is especially well-suited for the production of fatty acids. Moreover, it has a variety of genetic engineering tools and novel metabolic engineering strategies that make it a robust workhorse for the production of an array of value-added products. In this review, we summarize recent advances in metabolic engineering strategies for accumulating nutritional fatty acids in Y. lipolytica, including conjugated fatty acids and polyunsaturated fatty acids. In addition, the future prospects of nutritional fatty acid production using the Y. lipolytica platform are discussed in light of the current progress, challenges, and trends in this field. Finally, guidelines for future studies are also emphasized.