Increased biomass and lipid production by continuous cultivation of Nannochloropsis salina transformant overexpressing a bHLH transcription factor

• Published in: Biotechnology and bioengineering Vol. 116; no. 3; pp. 555 - 568
• Main Authors: Kang, Nam Kyu, Kim, Eun Kyung, Sung, Min‐Gyu, Kim, Young Uk, Jeong, Byeong‐ryool, Chang, Yong Keun
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01.03.2019; John Wiley and Sons Inc
• Subjects: Algae; Basic Helix-Loop-Helix Transcription Factors - genetics; Basic Helix-Loop-Helix Transcription Factors - metabolism; Batch Cell Culture Techniques - methods; Batch culture; bHLH transcription factor; Binding sites; Biofuels; Biomass; Biomaterials; Biomedical materials; continuous cultivation; Continuous culture; Cultivation; Culture; Dilution; Fatty acids; Fatty Acids - metabolism; Genes; Genetic Engineering; Helix-loop-helix proteins (basic); Lipid metabolism; Lipids; Metabolism; microalgae; Microalgae - genetics; Microalgae - metabolism; Nannochloropsis salina; Nitrogen; Nutrients; Phenotypes; Polymerase chain reaction; Stramenopiles - genetics; Stramenopiles - metabolism; Transcription factors; microalgae; biofuels; continuous cultivation; bHLH transcription factor; nannochloropsis salina
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.26894

Microalgae are promising feedstocks for sustainable and eco‐friendly production of biomaterials, which can be improved by genetic engineering. It is also necessary to optimize the processes to produce biomaterials from engineered microalgae. We previously reported that genetic improvements of an industrial microalga Nannochloropsis salina by overexpressing a basic helix‐loop‐helix transcription factor (NsbHLH2). These transformants showed an improved growth and lipid production particularly during the early phase of culture under batch culture. However, they had faster uptake of nutrients, resulting in earlier starvation and reduced growth during the later stages. We attempted to optimize the growth and lipid production by growing one of the transformants in continuous culture with variable dilution rate and feed nitrogen concentration. Relative to wild‐type, NsbHLH2 transformant consumed more nitrate at a high dilution rate (0.5 day −1), and had greater biomass production. Subsequently, nitrogen limitation at continuous cultivation led to an increased fatty acid methyl ester production by 83.6 mg l −1 day −1. To elucidate genetic mechanisms, we identified the genes containing E‐boxes, known as binding sites for bHLH transcription factors. Among these, we selected 18 genes involved in the growth and lipid metabolism, and revealed their positive contribution to the phenotypes via quantitative real‐time polymerase chain reaction. These results provide proof‐of‐concept that NsbHLH2 can be used to produce biomass and lipids. Overexpression of NsbHLH2 in Nannochloropsison salina altered expression of genes related to carbon metabolism, resulting in increased biomass and lipid production. However, the performance became similar to WT at the end of cultivation due to nutrient depletion under batch cultivation. In this study, we thus performed continuous cultivation for the transformant to accommodate its faster nutrient absorption, leading to optimized productivities of biomass and lipids. As a result, we significantly increased lipid productivity of the NsbHLH2 transformant.