An Escherichia coli Strain with Improved Itaconic Acid Stress Resistance
A technology of Escherichia coli and dry bacteria, applied in the field of microbial engineering, can solve the problems of accumulation of by-products, weakening of existing metabolic pathways, osmotic stress, etc.
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Embodiment 1
[0031] Example 1: Adaptive evolution of E. coli tolerant to higher itaconic acid concentrations
[0032] Taking strain E.coli K12 MG1655 as the starting strain, three parallel samples were made at the same time, and cultured in M9 medium at 37°C for 5 hours to the mid-logarithmic phase of bacterial growth; the strains after culturing for 5 hours were transformed with 4% of the inoculation volume. Connected to M9 medium containing 2.4g / L itaconic acid, placed in a shaker at 37°C at 220 rpm and cultured to an OD of 0.9-1.1, observe and record the growth time to reach the OD value; inoculate again with 4% The volume ratio of the strains cultivated to OD of 0.9-1.1 was transferred to the M9 medium of the same itaconic acid concentration, and the transfer was continued in turn, until the growth rate at the itaconic acid concentration was the same as that in normal without itaconic acid. Under the same or similar conditions of M9 medium, transfer to the next concentration, and so on...
Embodiment 2
[0033] Example 2: Growth of acclimated strains and starting strains under normal conditions
[0034] Specific steps are as follows:
[0035] (1) The strain E.coli K12 MG1655 (Control) and the strains E.coli I2, E.coli I3 and E.coli I4 obtained in Example 1 were respectively inoculated into LB liquid medium for activation, and placed at 37°C Incubate overnight at 220 rpm in a shaker;
[0036] (2) transfer the seed liquid obtained in the above (1) into LB liquid medium with an inoculation amount of 2% (V / V) respectively, and place it in a 37° C. shaker for cultivation at 220 rpm;
[0037] (3) in the culturing process of (2), sampling every 2 hours, measure the OD value under the wavelength of 600nm, draw the growth curve (the growth curve obtained by drawing is as follows: figure 1 ).
[0038] The result is as figure 1 As shown, through the growth performance test analysis, compared with the control, the growth curves of E.coli I2, E.coli I3 and E.coli I4 can basically keep ...
Embodiment 3
[0039] Example 3: Growth of the domesticated strain and the starting strain under the initial stress condition of 3.4 g / L itaconic acid
[0040] Specific steps are as follows:
[0041] (1) The strain E.coli K12 MG1655 (Control) and the strains E.coli I2, E.coli I3 and E.coli I4 obtained in Example 1 were respectively inoculated into M9 liquid medium for activation, and placed at 37°C Incubate overnight at 220 rpm in a shaker;
[0042] (2) The seed solution obtained in the above (1) was transferred to the M9 liquid medium containing 2.4g / L itaconic acid with the inoculum amount of 2% (V / V) respectively, and placed in a shaker at 37°C. 220rpm culture;
[0043](3) in the culturing process of (2), sampling every 2 hours, measure the OD value under the wavelength of 600nm, draw the growth curve (the growth curve obtained by drawing is as follows: figure 2 ).
[0044] The result is as figure 2 As shown, through the analysis of growth performance test, after culturing for 18h,...
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