Mechanisms On Improving Photosynthetic Characterization And Carbon Fixation Rate Of Microalgae By Nuclear Radiation And CO₂ Stress

Microalgae have been a research hotspot in the field of carbon sequestration and sustainable energy given their incomparable advantages,such as high efficiency for solar energy utilization,grow extremely fast and their high-valued biomass.In this study,three superior microalgae species Chlorella,Arthrospira and Nannochloropsis were mutated by nuclear radiation and domesticated under high CO2 concentration in flue gas to improve their growth and carbon fixation rates.Chlorophyll a variable fluorescence,flash oxygen and high-throughput transcriptome sequencing were applied on the analysis of carbon fixation system and metabolic network,revealing the reaction mechanisms on the enhanced photosynthetic characterization and carbon fixation process of microaglae.The biomass yield of Arthrospira mutant ZJU9000 was 176%higher than that of wild type on day 4.The results of transcriptome sequencing showed that the efficiency of photosynthesis was enhanced by the elevated levels of chlorophyll a and carotene.The increased biosynthesis rates of ribose phosphate and nucleotides increased the production of genetic materials for cell proliferation.Glycolysis and TCA cycle were enhanced to provide more energy material ATP and[H].Vitamins syntheses were enhanced and promoted cell growth.Furthermore,the carbon concentration mechanism was enhanced,and increased the utilization efficiency of CO2 from air.These improvements contributed to the enhanced carbon fixation ability in Arthrospira mutant ZJU9000.The biomass productivity and lipid content of Arthrospira mutant ZJU9000 were increased by 37.9%and 32.5%after domestication under 15 vol.%CO2.Gene sequencing results revealed that proton transport from the matrix to the intermembrane was enhanced,subsequently increasing the potential energy among the membranes;Meanwhile,the up-regulation of ATP synthase catalyzed the thesis of ATP with higher efficiency.More NADH was provided through tricarboxylic acid cycle.The amounts of the synthesised vitamins K1 and K2 were increased,which promoted electron transport during the photoreaction and release the free energy to transport protons and generate high potential energy.These processes resulted in more carbohydrates and lipids synthesized with more energy,besides,cell proliferation was accelerated and increased the biomass yield of Arthrospira.In the fatty acid synthesis metabolic pathway,more pyruvate flowed into malonyl-CoA through acetyl-CoA and entered into fatty acid synthesis.Concentration of H+ increased when CO2 was bubbling into the culture,thus the hydrogenation of carbon chains was enhanced and promoted lipid accumulation.Nannochloropsis oceanica was mutated with nuclear radiation to obtain improved phenotype with enhanced photosynthesis.Nannochloropsis mutant ZJU700 had a 30.2%higher oxygen evolution rate than wild type cells.High-throughput transcriptome sequencing showed that syntheses of antenna proteins were enhanced and increased solar utilization efficiency,photoreactions relevant genes were comprehensively up-regulated and generated more energy metarial ATP for dark reaction.CO2 fixation process was enhanced in dark reaction and more carbohydrates were synthesized.Photosynthesis efficiency was further improved as more Vitamin E and Vitamin K1,K2 were generated and promoted the electron transport.Chlorella sp.,which was adapted to seawater,was mutated with nuclear irradiation and biomass yield of the Chlorella mutant MS700 increased by 25%.Chlorophyll-a variable fluorescence and oxygen evolution proved that,chlorophyll-based flash oxygen yield was 104.2%higher,higher photochemical efficiency Fv/Fm value showed higher utilization efficiency of electrons generated in photosystem Ⅱ,Chlorella mutant had a 27.7%higher capability of incorporating its dissolved inorganic carbon.These results consistently demonstrated the effectiveness of mutant MS700 in carbon fixation and higher biomass yield.Medium of Chlorella mutant was optimized to improve carbon fixation rate and lipid productivity of Chlorella.TEM results showed that,cell number increased as cell proliferation accelerated when initial nitrogen and phosphorus concentration increased,whereas cell diameter and cell wall thickness decreased by 26.6%and 69.7%,respectively.The long-chain unsaturated fatty acids increased and short-chain saturated fatty acids decreased.The growth rate of Chlorella mutant was higher than wild type Chlorella,thus the cell wall thickness was 36.2%thinner and direct lipid extraction rate increased by 33%.Pilot scale tests were carried out in Arthrospira mutant ZJU9000 and Chlorella mutant MS700,the biomass yields of these two mutants in raceway ponds were 29.9%and 25%higher,respectively,than their corresponding wild type.These results provided technical support for the large-scale industrial application of carbon sequestration by microalgae.