Stable transformation of the green algae Acutodesmus obliquus and Neochloris oleoabundans based on E. coli conjugation

Camilo F Muñoz,Mark H.J Sturme,Sarah D'Adamo,Ruud A Weusthuis,René H Wijffels

doi:10.1016/j.algal.2019.101453

Camilo F Muñoz, Mark H.J Sturme + Show 3 more

Open Access

https://doi.org/10.1016/j.algal.2019.101453

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Abstract

Microalgae are an ideal platform for the production of high-value chemicals, nutritional products and biofuels. Genetic engineering could speed up the development of microalgae derived products and reduce the overall production costs. Genetic methods such as particle bombardment, electroporation, Agrobacterium tumefaciens mediated transformation (ATMT), and agitation with glass beads and silicon carbide whiskers have been developed for the genetic transformation of microalgae. However, the transformation efficiency is species dependent, so a variety of transformation methods are required to engineer a wide range of microalgae species. The oleaginous microalgae Acutodesmus obliquus and Neochloris oleoabundans have a great potential as production platforms due to their ability to produce large amounts of triacylglycerol (TAG). Genetic modification techniques however are required to increase TAG levels further or to modify the fatty acid composition. Recently, a conjugation-based method for the delivery of episomes from bacteria to diatom microalgae has been reported. In this study, we have achieved the successful transformation of green oleaginous microalgal strains by transferring an expression vector via conjugation from E. coli. Since delivery of exogenous DNA into the microalgae cells is only the first step in obtaining transgenic microalgae, we further analyzed transformation efficiencies by PCR and expression of the Clover fluorescent protein in the targeted species.

Highlights

Microalgae are considered as promising feedstock for the production of numerous valuable commercial products which can be used for the production of biofuels, cosmetics, food and feed [1,2,3,4,5,6,7]
We report for the first time an efficient and stable transformation of the green microalgae Acutodesmus obliquus and Neochloris oleoabundans by transferring exogenous DNA from Escherichia coli via conjugation
We evaluated the effect of hygromycin B, paromomycin and zeocin on the cell growth of A. obliquus and N. oleoabundans

Summary

Introduction

Microalgae are considered as promising feedstock for the production of numerous valuable commercial products which can be used for the production of biofuels, cosmetics, food and feed [1,2,3,4,5,6,7]. Among transformation methods for the delivery of exogenous DNA the most common techniques are electroporation, Agrobacterium tumefaciensmediated transformation (ATMT), ballistic systems and agitation with glass beads [10,11,12,13]. Most of these techniques have been proven to work with great success in model strains such as Chlamydomonas reinhardtii, Phaeodactylum tricornutum, and Chlorella sp., there is a lack of efficient and stable transformation techniques that can be applied to a broader range of microalgae strains

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