| This study included two parts: the first part involved in the comparation and optimization of media tested. From five media(YMA, TY, PA, SM and BSE), choose a low cost medium suitable for the strains tested. The strains tested were Sinorhizobiumfredii, HN01 and Bradyrhizobium japonicum, USDA110. The optical density of the strains tested was tested by UV7501 spectrophotometer and there OD growth curves were made. The results showed that the growth speed of HN01 was similar in media TY, YMA and BSE, which was faster than in PA and SM. The growth speed of USDA110 was fast in medium BSE, followed by TY and YMA, slowest in SM.the growth curve of HN01 and USDA110 growing respectively in TY and BSE was also tested. The results showed the generation time of HN01 in TY was 3.3 hours and in BSE was 2.4 hours. The generation time of USDA110 in TY was 7.8 hours and in BSE was 6 hours. BSE was suitable to be used for industrial fermentation.Since growth speed of USDA110 was very slow in YMA, the components of media were optimized to find a medium suitable for USDA110. At first, the growth speed of USDA110 in different carbon sources was compared, the results showed that the growth speed in glucose was fast, after fifty-hours cultivation, the optical density of USDA110 in glucose was 1.05 and only 0.68 in mannitol. the growth speed is slowest in lactose. Then glucose was chosen as carbon source to do the L,9(34) orthogonal optimization experiment. The results showed that the third medium was best. Except the seventh medium, all the other media tested could increase growth speedofUSDAHO.The results from statistical analysis showed the glucose and the yeast extract played highly significant effect on the growth speed, while K2HPO4 and MgSO4-7H2O showed little effect on the growth speed. The optimum medium components were glucose 15g, yeast extract 4g , K2HPO4 0.5g, MgSO4-7H2O 0.2g, NaCl 0.1 g.CaCl2 0.05g, Rh trace solution 4ml and dH2O 996ml. The following experiment showed that the growth speed of USDA110 in optimum medium was similar to that in the third medium.The second part involved in the production of rhizobium inoculant. The viable ratio of different solid, liquid and frozen inoculants was compared during different storage time. The solid peat, vermiculite and perlite inoculants were stored at room temperature and sampled at regulate time to count the viable number. The results . showed the viable number of HN01 in peat inoculant was 4.5 times more in the first fifteen days, then began to reduce, after a month, the viable number reduced to thestart level. At first three months, the viable number was high, but after that the death ratio increased sharply. The growth speed of USDA110 in peat inoculant was slow, but increase time was longer than that of HN01, the viable number was 1.5 times more in the first month, The viable number reduced to the start lever after two months,, and the death ratio was also slower than that of HN01. Among the three carriers tested, peat was better than vermiculite, the latter was better than perlite.Liquid concentratiod inoculant was prepared by centrifugation, The results showed both the viable number and viable ratio of the strains tested were high. After six months, there were still 1 billion viable number of USDA110 left, and 2 billion viable number of HN01 left. The viable ratio of HN01 in liquid 1 and liquid 2 was respectively 9.4%, 9.6% after four months, the viable ratio of HN01 in liquid 1 and liquid 2 reduced respectively to 5%, 5.4% after five months, The viable ratio of USDA110 in liquid 1 and liquid 2 was respectively 12.2%, 11.7% after four months, The viable ratio of USDA110 both in liquid 1 and liquid 2 reduced approximately to 4% after five months. It was better for liquid inoculant to be stored for three or four months. The viable ratio of liquid inoculant kept in nitrogen gas and in vacuum was similar. The results of freeze-dry inoculant showed the viable ratio of the strains tested was the highest during storaged at -20℃, followed by the storaged at 4?... |
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