Enhanced Lipid Productivity and Photosynthesis Efficiency in a <i>Desmodesmus</i> sp. Mutant Induced by Heavy Carbon Ions

<div><p>The unicellular green microalga <i>Desmodesmus s</i>p. S1 can produce more than 50% total lipid of cell dry weight under high light and nitrogen-limitation conditions. After irradiation by heavy ¹²C⁶⁺ ion beam of 10, 30, 60, 90 or 120 Gy, followed by screening of resulting mutants on 24-well microplates, more than 500 mutants were obtained. One of those, named D90G-19, exhibited lipid productivity of 0.298 g L⁻¹⋅d⁻¹, 20.6% higher than wild type, likely owing to an improved maximum quantum efficiency (Fv/Fm) of photosynthesis under stress. This work demonstrated that heavy-ion irradiation combined with high-throughput screening is an effective means for trait improvement. The resulting mutant D90G-19 may be used for enhanced lipid production.</p></div

Composition of total lipid in the <i>Desmodesmus</i> sp.

a<p>Microalgae cells were harvested after grown under HL-N (300–400 µmol photons m⁻²⋅s⁻¹, nitrogen-depleted BG-11 medium with 4.25 mM NaNO₃) for 8 days. Mean ± SE with three replicates.</p>b<p>G:P: ratio of glycolipid to phospholipid.</p

Total lipid contents of <i>Desmodesmus</i> sp.

<p>WT and D90G-19 were grown under HL-N conditions (300–400 µmol photons m⁻²⋅s⁻¹, nitrogen-limited BG-11 medium with 4.25 mM NaNO₃). The total lipid of the mutant D90G-19 was significantly higher than WT at day 4 (P = 0.0051), day 6 (P = 0.006) and day 8 (P = 0.0039). D90G-19: closed square; WT: open square.</p

The consumption of NaNO₃ and growth kinetics of <i>Desmodesmus</i> sp.

<p>(A) The consumption of NaNO₃ in the nitrogen-limited (4.25 mM) BG-11 medium (closed symbols) and growth kinetics (open symbols) of <i>Desmodesmus</i> sp. S1 under low light (closed square, open triangle) and high light (closed circle, open diamond), and (B) biomass concentration of <i>Desmodesmus</i> sp. S1 wild type (WT, open square) and mutant D90G-19 (closed square) cultivated in BG-11 medium under LL+N conditions (50 µmol photons m⁻²⋅s⁻¹, BG-11 medium with 17 mM NaNO₃).</p

Photosynthesis efficiency of <i>Desmodesmus</i> sp. (A) Potential maximum quantum efficiency (Fv/Fm) and (B) oxygen evolution rate.

<p>WT and D90G-19 were grown under HL-N (300–400 µmol photons m⁻²⋅s⁻¹, nitrogen-limited BG-11 medium with 4.25 mM NaNO₃). D90G-19: closed circle; WT: open square.</p

Fatty acid compositions of individual lipid classes in <i>Desmodesmus</i> sp.^d.

a<p>percentage of unsaturated fatty acids in total fatty acids.</p>b<p>the degree of fatty acid unsaturation <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060700#pone.0060700-Chen2" target="_blank">[41]</a>.</p>c<p>not detected.</p>d<p>Microalgae were harvested after grown under HL-N (300–400 µmol photons m⁻²⋅s⁻¹, nitrogen-depleted BG-11 medium with 4.25 mM NaNO₃) for 8 days. Mean ± SE with three replicates.</p

Starch contents of <i>Desmodesmus</i> sp. WT and D90G-19.

<p>The initial starch content was measured in algae cells grown under LL+N for 4 days. Afterwards, the cells were grown under HL-N (300–400 µmol photons m⁻²⋅s⁻¹, nitrogen-limited BG-11 medium with 4.25 mM NaNO₃). D90G-19: closed square; WT: open square.</p

Relative amount of lutein in the progenies and in the control group during HL exposure.

<p>The concentration of lutein was measured by HPLC. The five progenies and the control group were grown under HL exposure (300–400 μmol photons m^-2 s^-1) for 96 h, and each value obtained during HL exposure was compared with that obtained under normal conditions (0 h). n = 3+SD.</p

Fv/Fm of PSII of the progenies and of the control group.

<p>The Fv/Fm of the control group and of the progenies was determined at the indicated time points after dark adaptation for 8 min. Minimal fluorescence in the dark and maximal fluorescence upon a saturating light pulse were measured by Maxi-IMAGING-PAM at room temperature. The initial inoculation amount of each progeny and of the control group was 6.25% by volume; the Fv/Fm of the initial algal cell cultures (0 d) was too low to measure. n = 3+SD.</p

Relative amount of DES in the progenies and in the control group under HL exposure.

<p>The concentrations of Vx, Ax and Zx were measured by HPLC. <i>DES</i> = (<i>Zx</i>+0.5<i>Ax</i>)/(<i>Vx</i>+<i>Ax</i>+<i>Zx</i>). The five progenies and the control group were grown under HL exposure (300–400 μmol photons m^-2 s^-1) for 96 h. Each experimental HL exposure value was compared with the value found under normal conditions (0 h). n = 3+SD.</p