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Lipid production in Nannochloropsis gaditana is doubled by decreasing expression of a single transcriptional regulator

Nature Biotechnology volume 35, pages 647652 (2017) | Download Citation

Abstract

Lipid production in the industrial microalga Nannochloropsis gaditana exceeds that of model algal species and can be maximized by nutrient starvation in batch culture. However, starvation halts growth, thereby decreasing productivity. Efforts to engineer N. gaditana strains that can accumulate biomass and overproduce lipids have previously met with little success. We identified 20 transcription factors as putative negative regulators of lipid production by using RNA-seq analysis of N. gaditana during nitrogen deprivation. Application of a CRISPR–Cas9 reverse-genetics pipeline enabled insertional mutagenesis of 18 of these 20 transcription factors. Knocking out a homolog of fungal Zn(II)2Cys6-encoding genes improved partitioning of total carbon to lipids from 20% (wild type) to 40–55% (mutant) in nutrient-replete conditions. Knockout mutants grew poorly, but attenuation of Zn(II)2Cys6 expression yielded strains producing twice as much lipid (5.0 g m−2 d−1) as that in the wild type (2.5 g m−2 d−1) under semicontinuous growth conditions and had little effect on growth.

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Acknowledgements

This work was funded by ExxonMobil and Synthetic Genomics, Inc. We thank A. Withrow (Center for Advanced Microscopy at Michigan State University) for producing the TEM images and C. Packard, B. Scherer, E. Wang and the rest of the analytical team at SGI for processing FAME and TOC samples. This work is dedicated to our colleague Tom Carlson, who passed away during the preparation of this manuscript.

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    • Tom J Carlson

    Deceased.

Affiliations

  1. Synthetic Genomics Inc., La Jolla, California, USA.

    • Imad Ajjawi
    • , John Verruto
    • , Moena Aqui
    • , Leah B Soriaga
    • , Jennifer Coppersmith
    • , Kathleen Kwok
    • , Luke Peach
    • , Elizabeth Orchard
    • , Ryan Kalb
    • , Weidong Xu
    • , Tom J Carlson
    • , Kristie Francis
    • , Katie Konigsfeld
    • , Judit Bartalis
    • , Andrew Schultz
    • , William Lambert
    • , Ariel S Schwartz
    • , Robert Brown
    •  & Eric R Moellering

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Contributions

I.A. and E.R.M. conceived the study and designed experiments. R.B. provided technical advice. E.O. and R.K. designed the productivity assays. L.B.S. and A.S.S. performed computational and bioinformatics analyses. M.A., J.V., J.C., L.P., J.B., A.S., W.X., T.J.C., K.F., W.L., K. Kwok and K. Konigsfeld performed the experiments. I.A. and E.R.M. wrote the manuscript with support from all authors.

Competing interests

I.A., J.V., M.A., J.C., K. Kwok, L.P., E.O., R.K., K. Konigsfeld, A.S., W.L., R.B. and E.R.M. are employees of Synthetic Genomics, Inc. Synthetic genomics has filed patents related to this work, with I.A., J.V., M.A., L.B.S. and E.R.M. listed as inventors.

Corresponding authors

Correspondence to Imad Ajjawi or Eric R Moellering.

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https://doi.org/10.1038/nbt.3865