Development of a multigene expression system using 2A peptides in Rhodosporidium toruloides

IF 3.5 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biotechnology and Bioengineering Pub Date : 2024-09-17 DOI:10.1002/bit.28843

Xiao Guo, Zhenzhen Bai, Huimin Zhao, Shuobo Shi

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Abstract

In eukaryotes, gene expression typically requires individual promoter and terminator for each gene, making the expression of multiple genes tedious and sometimes too difficult to handle. This is especially true for underdeveloped nonmodel organisms with few genetic engineering tools and genetic elements such as Rhodosporidium toruloides. In contrast, polycistronic expression offers advantages such as smaller size and ease of cloning. Here we report the development of a multigene expression system using 2A peptides in R. toruloides. First, twenty‐two 2A peptides were evaluated for their cleavage efficiencies, which ranged from 33.65% to 93.32%. Subsequently, the 2A peptide of ERBV‐1 with the highest efficiency was selected to enable simultaneous expression of four proteins. In addition, we demonstrated the optimization of the α‐linolenic acid biosynthetic pathway using ERBV‐1 peptide mediated polycistronic expression, which increased the α‐linolenic acid production by 104.72%. These results suggest that using ERBV‐1 peptide is an efficient strategy for multigene expression in R. toruloides.

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利用 Toruloides 罗多孢子虫中的 2A 肽开发多基因表达系统

在真核生物中，基因表达通常需要为每个基因设置单独的启动子和终止子，这使得多基因表达变得繁琐，有时甚至难以处理。这对于基因工程工具和遗传因子较少的不发达非模式生物（如红孢子虫）来说尤其如此。相比之下，多核苷酸表达具有体积小、易于克隆等优点。在此，我们报告了利用 2A 肽在环孢子虫中开发多基因表达系统的情况。首先，我们评估了 22 种 2A 肽的裂解效率，其范围从 33.65% 到 93.32%。随后，我们选择了效率最高的 ERBV-1 的 2A 肽，以实现四种蛋白质的同时表达。此外，我们还证明了利用ERBV-1多肽介导的多聚体表达优化了α-亚麻酸的生物合成途径，使α-亚麻酸的产量提高了104.72%。这些结果表明，使用ERBV-1多肽是在R.

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来源期刊

Biotechnology and Bioengineering 工程技术-生物工程与应用微生物

CiteScore

7.90

自引率

5.30%

发文量

280

审稿时长

2.1 months

期刊介绍： Biotechnology & Bioengineering publishes Perspectives, Articles, Reviews, Mini-Reviews, and Communications to the Editor that embrace all aspects of biotechnology. These include: -Enzyme systems and their applications, including enzyme reactors, purification, and applied aspects of protein engineering -Animal-cell biotechnology, including media development -Applied aspects of cellular physiology, metabolism, and energetics -Biocatalysis and applied enzymology, including enzyme reactors, protein engineering, and nanobiotechnology -Biothermodynamics -Biofuels, including biomass and renewable resource engineering -Biomaterials, including delivery systems and materials for tissue engineering -Bioprocess engineering, including kinetics and modeling of biological systems, transport phenomena in bioreactors, bioreactor design, monitoring, and control -Biosensors and instrumentation -Computational and systems biology, including bioinformatics and genomic/proteomic studies -Environmental biotechnology, including biofilms, algal systems, and bioremediation -Metabolic and cellular engineering -Plant-cell biotechnology -Spectroscopic and other analytical techniques for biotechnological applications -Synthetic biology -Tissue engineering, stem-cell bioengineering, regenerative medicine, gene therapy and delivery systems The editors will consider papers for publication based on novelty, their immediate or future impact on biotechnological processes, and their contribution to the advancement of biochemical engineering science. Submission of papers dealing with routine aspects of bioprocessing, description of established equipment, and routine applications of established methodologies (e.g., control strategies, modeling, experimental methods) is discouraged. Theoretical papers will be judged based on the novelty of the approach and their potential impact, or on their novel capability to predict and elucidate experimental observations.

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