ARTP Mutagenesis of Schizochytrium sp. PKU#Mn4 and Clethodim-Based Mutant Screening for Enhanced Docosahexaenoic Acid Accumulation.

Jiantao Li,Lu Liu,Xianhua Liu,Biswarup Sen,Guangyi Wang,Sai Zhang,Mohan Bai

doi:10.3390/md19100564

Abstract

Schizochytrium species are one of the best oleaginous thraustochytrids for high-yield production of docosahexaenoic acid (DHA, 22:6). However, the DHA yields from most wild-type (WT) strains of Schizochytrium are unsatisfactory for large-scale production. In this study, we applied the atmospheric and room-temperature plasma (ARTP) tool to obtain the mutant library of a previously isolated strain of Schizochytrium (i.e., PKU#Mn4). Two rounds of ARTP mutagenesis coupled with the acetyl-CoA carboxylase (ACCase) inhibitor (clethodim)-based screening yielded the mutant A78 that not only displayed better growth, glucose uptake and ACCase activity, but also increased (54.1%) DHA content than that of the WT strain. Subsequent optimization of medium components and supplementation improved the DHA content by 75.5 and 37.2%, respectively, compared with that of mutant A78 cultivated in the unoptimized medium. Interestingly, the ACCase activity of mutant A78 in a medium supplemented with biotin, citric acid or sodium citrate was significantly greater than that in a medium without supplementation. This study provides an effective bioengineering approach for improving the DHA accumulation in oleaginous microbes.

Highlights

This study provides the first report of a bioengineering approach, integrating ARTP mutagenesis and clethodim-based screening strategy, which yielded a thraustochytrid mutant with a better capacity for DHA accumulation than its parental strain
Like other conventional mutagenesis methods, the treatment duration can be critical for achieving the desirable mutation efficiency [42]
PKU#Mn4 was found to be sensitive to the ARTP exposure time (Figure 1a)

Summary

Introduction

As one of the most important polyunsaturated fatty acids (PUFA), docosahexaenoic acid (DHA, 22:6) benefits the development of the fetus’s nerves and retina and prevents several human diseases such as cardiovascular disease, hypertension and thrombi [1]. Given the multiple benefits to human health, DHA has been an important functional food component with high demand on the global market. As the current major source of DHA, deep-sea fishes obtain PUFA from marine microorganisms, marine microalgae and protists [2,3,4]. The supply of fish oil is limited by food security, environmental and toxicity concerns and, further research is needed to develop efficient and sustainable alternative sources of DHA [5]

Methods

Results

Conclusion