Studies On Photosynthesis And Oil Accumulation Of Epiphytic/Endophytic Microalgae Of Aconitum

Microalgae,on the one hand,are rich in nutrients and high value bioactive substance,on the other hand,they play a special role in biological evolution and ecosystem.Heveochlorella roystonensis and H.hainanensis were previously isolated and identified as two closely related species,and were studied in this research,focusing on their chlorophyll,lipid,fatty acid composition and extracellular secretion.The differences of their phenotypic changes and the transcriptional regulation of photosynthesis and lipid metabolism of the two microalgae was also studied.The results lay a foundation for the understanding the evolution of microalgae from epiphytic to endophytic,from autotrophic to heterotrophic,and for the comprehensive utilization of microalgae.The main achievements are summarized as follows:(1)The optimal sucrose concentration,growth curve,biomass and p H value of culture liquid of the two species were studied.Results showed that: under the condition of no carbon source,the growth of H.roystonensis was normal with a biomass of 3.20 g/ L,while the growth of H.hainangensis was slow with a biomass ofonly 0.09 g / L;0.5% sucrose was the most suitable carbon concentration for the growth of H.hainangensis,the biomass was increased to 3.87 g/L.The p H value of the culture medium increased to be alkaline by culturing H.roystonensis,while the p H value decreased to be acidic by culturing H.hainangensis.With the addition of carbon source,the alkalinity of the culture medium of H.roystonensis increased and the acidity of the culture medium of H.hainangensis decreased significantly,the latter was as low as 4.7.(2)The optimum chlorophyll extraction condition was 85% of ethanol andextracting for 15 min at 45 ℃.The chlorophyll a,b and total chlorophyll contents of H.roystonenesis were significantly higher than that of H.hainangensis,and were increased by light treatment.The chlorophyll b and total chlorophyll content of H.hainangensis were increased,but the chlorophyll a content was decreased by light treatment;when the carbon source was added,the chlorophyll a content of the two strains was decreased,while the chlorophyll b and total chlorophyll contentswere increased.Taken together,the photosynthesis efficiency of H.roystonensis was significantly higher than that of H.hainangensis.(3)The total oil content of H.hainangensis was significantly higher than that of H.roystonensis.The oil content of H.hainangensis was 30.65% after 15 days of dark culture with carbon source,which was 5.4 times of that of H.roystonensis after 10 days of light culture without carbon source.The oil accumulation of H.hainangensis was significantly increased by 27.15-31.43%,but the effect on H.roystonensis was not obvious.The content of fatty acids in H.hainangensis is relatively single,mainly composed of 7 fatty acids.When carbon source was added,the saturated fatty acid content of H.roystonensis decreased,while that of H.hainangensis was to the oposite.(4)A total of 140 extracellularmetabolites were identified in the culture media of H.roystonensis and H.hainangensis,among which the differently presented metabolites were carboxylic acids and pantothenic acids.When carbon source was supplied,the content of vitamin A acid was the highest,accounting for 61.73% and 23.59% of the organic acids in H.hainangensis and H.roystonensis,respectively.With the addition of carbon source,the contents of organic acids increased,while the contents of amines and alkaloids decreased,which was consistent with the pattern of p H value of the two species.(5)The chloroplast genome of H.hainangensis was 2636877 bp,with 122 genes,while the chloroplast genome of H.roystonensis contains 216 879 bp with 118 genes.However,the genes controlling photosynthesis of H.roystonensis were more than those of H.hainangensis.The chloroplast memberane protein cem A has long additional amino acid residues in the variable region compared to that of H.roystonensis,which may have a negative influence on its photosynthesis.Another gene acc D in H.hainangensis has a length of twice as long as that of H.roystonenesis,which may increase its ability of lipid biosynthesis..(6)Five differentially expressed genes related to oil biosynthesis were identified in H.hainangensis,while only two were identified in H.roystonensis,,when carbon source was present or absent;and12 differentially expressed genes were identified between the two species,of which 6 were up-regulated and 6 were down regulated.Between light and dark treatments,there were 54 differentially expressed genes regulating photosynthesis in H.roystonensi,of which 31 were up-regulated by light,while 32 genes were differentially expressed in H.hainangensis,of which only 3were up-regulated.Thirty differentially expressed genes related to photosynthesis,lipid metabolism and energy metabolism were selected and verified by q PCR.Seven key differentially expressed genes were confirmed,among which AK,ATP like,acyl-Co A and apt were involved in lipid metabolism,and Qxygen protein,Psb P and PP2 C were involved in photosynthesis.