Enterococcus faecium strain capable of producing L-lactic acid
A technology of Enterococcus faecium and strains, applied in the field of Enterococcus faecium strains, can solve the problems of insufficient research and development of L-lactic acid, etc., and achieve the effect of high conversion rate
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Embodiment 1
[0045] Example 1: Screening of Enterococcus faecium strains producing L-lactic acid
[0046] Take samples such as soil from animal feces, rumen contents of cattle and sheep according to conventional methods, take 2g samples under aseptic conditions, put them into a 250mL triangular flask filled with 40mL sterile water, take 1mL after shaking and inoculate into the container Incubate in a 250mL Erlenmeyer flask with 40mL enrichment medium at 37°C and 180r / min shaker for 24h, then dilute the enrichment solution onto bromocresol purple-calcium carbonate screening plate, and keep the temperature in an anaerobic bag at 37°C Cultivate, and after a single colony grows, pick a colony with a large color-changing transparent circle and puncture it for preservation. Then these preserved strains were screened by shake flask fermentation, 40mL shake flask fermentation medium was placed in a 250mL Erlenmeyer flask, one branch was connected to one bottle with 1-2 rings of slant strains, 37°C...
Embodiment 2
[0047] Embodiment 2: the breeding of L-lactic acid faecium Enterococcus
[0048] Inoculate the liquid seed medium with the starting strain Enterococcus faecium H-36, cultivate it to the middle logarithmic growth phase at 37°C, collect the bacteria by centrifugation, wash twice with sterile normal saline, and then add sterile normal saline to make a bacterial suspension. Keep the cell concentration at 1×108-109 cells / ml. Take 10ml of the above-mentioned bacterial suspension and put it into a plate, put it under a 30W ultraviolet lamp to irradiate, the irradiation distance is 15cm, and the irradiation time is 3 minutes, 5 minutes, 7 minutes, 10 minutes. Bromocresol Purple-Calcium Carbonate Screening Plate for Calcium Lactate, incubate at 37°C for 1-2 days in the dark, pick a single colony that grows well and has a large discoloration transparent circle for preservation, and culture at 37°C for 1-2 days, after the slant is mature Inoculate the shake flask with fermentation mediu...
Embodiment 3
[0051] Example 3: Breeding of high-purity L-lactic acid bacteria strain Enterococcus faecium CGMCC No.7274
[0052] The mutant strain H-U36 strain screened in Example 2 was inoculated into the liquid seed medium, cultivated at 37°C to the middle of the logarithmic growth phase, collected the bacteria by centrifugation, washed twice with sterile normal saline, and then added sterile normal saline to make Make a bacterial suspension so that the cell concentration is 1×108-109 cells / ml, and set aside.
[0053] Preparation of nitrosoguanidine treatment solution: Weigh 20mg of nitrosoguanidine, place it in a 100ml sterile Erlenmeyer flask, add 2ml of acetone to dissolve it, then add 18ml of Tris buffer (pH6.0, 0.5mol / L) and mix well ,spare.
[0054] Take 10ml of the above-mentioned nitrosoguanidine treatment solution, add 10ml of bacterial suspension, incubate and shake at 30°C for 50-60 minutes, and take samples every 10 minutes. After sampling, first dilute 1000 times to stop th...
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