A strain of Escherichia coli producing l-lysine and its application
A technology of Escherichia coli and lysine, applied in the field of bioengineering, can solve the problems of low effect
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Embodiment 1
[0033] Example 1: Getting of mutant strains
[0034] Escherichia coli LQL separated from the bread in 2017 is a strain, and screening is performed in accordance with the following method:
[0035] (1) Always pressure room temperature plasma (ARTP) mutagenesis
[0036] 1 Preparation of bacteria suspension
[0037] The strain of Escherichia coli LQL was taken into a triangular bottle of 30 ml / 500 ml seed medium, 37 ° C, reciprocating shaker 100 rpm for 8 h to the middle and late stage, and the bacterial liquid is separated Qing, the bacteria was washed 3 times with PBS solution, and then resuspended with saline, so that the bacterial suspension OD 562 Return to 1.0.
[0038] 2ARTP mutagenesis
[0039] 10 μL of the prepared bacterial suspension is uniformly applied to the sterilized cooled small iron sheet, and placed in the ARTP breeding machine, the mutagenesis power is 100W, the high purity helium air air volume is 10 SLM, the treatment distance is 2mm, and 0, 20, respectively, ...
Embodiment 2
[0046] Example 2: 7.5L fermenter in E. coli and FMME-lys LQL case of producing L- lysine fermentation
[0047] (1) Preparation of seed was
[0048] The original strain of E. coli and mutant E. coli LQL FMME-lys were crossed on a solid plate medium, 37 [deg.] C incubator for 2-3 days in culture, inoculated with a single colony picked needle streaked on a slant medium for culture expanded, and then cultured in an incubator at 37 ℃ 1 to 2 days, good long-picked from the slant culturing a ring taken into seed medium, 35 ~ 37 ℃, 100 ~ 130rpm reciprocating shaker for 12 ~ 18h.
[0049] (2) 7.5L fermentor culture
[0050] Step seed liquid (1) was prepared according to the 12% was inoculated into the fermentor, the fermentor initial fermentation volume of 2L, fermentation temperature 37 ℃, aeration rate 1 vvm, the initial stirring speed of 400 rpm, followed by stirring, and dissolved oxygen associated with the rotational speed of 10 to 20%, when the glucose concentration is less than 15g ...
Embodiment 3
[0052] Example 3: In 7.5L of fermentor nitrogen coli FMME-lys-lysine fermentation influence L-
[0053] Example 2 Step seed solution (1) prepared in accordance with the 10% obtained was inoculated into the fermentor, the fermentor initial fermentation volume of 2L, were added 3g / L of ammonium sulfate, potassium nitrate and urea, the fermentation temperature of 37 ℃, aeration rate 1 vvm, stirring speed of 400 rpm initially, and then the stirring speed associated with dissolved oxygen controlled at 10 ~ 20%, when the glucose concentration is less than 15g / L, supplemented with glucose, the glucose concentration was maintained at 20 ~ 30g / L, when NH 4 + Concentration lower than 1g / L, supplemented with ammonium sulfate, of NH 4 + Concentration is maintained at 0.5-1g / L, taken during fermentation was centrifuged fermentation, L- lysine content was determined by ninhydrin color method supernatant was collected.
[0054] The results showed (see Table 1): ① When the initial nit...
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