The Breeding Resistance To High Glucose Concentrations And Optimization Of Culture Conditions Of Nostoc Flagelliforme Cells

Nostoc flagelliforme is a kind of biological nitrogen fixation activity of photoautotrophy terrestrial cyanobacterium. N. flagelliforme has high great economic and medicinal value. The mixotrophic modes was the optimal mode for obtaining the high cell density of N. flagelliforme, but the tolerance of glucose concentration for N. flagelliforme cells is lower. The wild N. flagelliforme was the starting strain in this paper, evolution engineering breeding was conducted with it and the strain that tolerance of high glucose concentration was screened. Then medium components and culture conditions were optimized with the selected N. flagelliforme cells. The main results were shown as follows:1. N. flagelliforme cells separated from field colony were chosen to as the starting strain. The Gene diversity library of it was established by UV-C mutagenic treatment. UV-B ultraviolet radiation and drought stress were the selection pressure in the evolutionary engineering breeding. Higher polysaccharide producing strain which having tolerance of high glucose concentration was screened. The optimal dosage of UV-C ultraviolet radiation for N. Flagelliforme cells was 24 h, and the optimal choice pressure of UV-B ultraviolet radiation and drought stress was 36h、2h. Respectively. The tolerance of glucose concentration of selected strain could reach 15 g/L.2. Plackett-Burman experimental design and response surface methodology were applied to screening and optimizing 6 factors for medium components and culture conditions of mixtrophic growth of N. flagelliforme cells, respectively. The optimal medium components for N. flagelliforme cells mixotrophic growth, discriminated from the regression equation are glucose 9.0 g/L, NaNO3 2.5 g/L and KH2PO4 0.31 g/L, and the yield biomass of N. flagelliforme cells is 2.96 g/L, which was increased 12.8% than before. The optimal culture conditions are culture temperature 25℃, light intensity 60 μmol. m-2 s-1 and medium initial pH 8.5, the cell dry weight of N. flagelliforme cells is 2.76 g/L, which was increased 27.4% than before.3. Light attenuation of N. flagelliforme cells in the mixotrophic and photoautotrophic cultivation was investigated. The adapted Lambert-Beer’s law could well simulate the influences of optical length and cell concentration on light attenuation, and the light attenuation equations of N. flagelliforme cells were established in photoautotrophic and mixotrophic cultivation, respectively. While the concentration of N. flagelliformecells density increased, light intensity obtained through N. flagelliforme cells culture declined. Especially, with the increase of optical length, the speed of light attenuation quickened evidently. When the biomass and incident light intensity were identical, the light attenuation inside photoautotrophic culture was faster than that in mixotrophic culture. For example, when the incident light intensity was 48.73 μmol.m-2.s-1, the cell dry weight of N. flagelliforme cells was 0.7715 g/L, the transmitted light intensity inside photoautotrophic culture and mixotrophic culture was 28.35 μmol.m-2.s-1 and 30.17 μmol.m-2.s"1 at optical length of 2 cm, respectively.The average carbon content of the N. flagelliforme cells was 40.75% according to the elemental analyzer. The photosynthetic efficiency was 14.3%.