A method for improving the biosynthesis of 1,3-propanediol
A technology of biosynthesis and propylene glycol, which is applied in the field of bioengineering, can solve the problems of no reports, etc., and achieve the effect of fast bacterial growth and high product concentration
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Embodiment 1
[0021] Example 1. Knocking out pck reduces the formation of the by-product 2,3-BD, and promotes the growth of bacteria and the synthesis of 1,3-PD
[0022] The M 2014574 strain was cultured overnight at 37° C. in LB medium (0.5% yeast extract, 1% tryptone, 1% NaCl, pH 7.0), and the genome was extracted. Using the extracted genome of M 2014574 as a template, primers were designed based on the pck gene (locus_tag: KPN_03773) on the genome sequence of Klebsiella pneumoniae MGH 78578 registered in NCBI (LOCUS: NC_009648), and the gel was recovered after the PCR reaction. The band was sequenced, and the gene analysis was performed on the target band after sequencing, and the similarity with the pck gene on MGH 78578 was 100%. The pck gene (1623bp) of the genome of M 2014574 was knocked out by means of homologous recombination, and the obtained recombinant strain was M 2014574△pck.
[0023] M 2014574 and M 2014574△pck were respectively inoculated in 250ml Erlenmeyer flasks for anae...
Embodiment 2
[0027] Example 2. Knocking out the ppc gene has almost no effect on the growth of bacteria and the synthesis of 2,3-butanediol and 1,3-PD
[0028]The M 2014574 strain was cultured overnight at 37° C. in LB medium (0.5% yeast extract, 1% tryptone, 1% NaCl, pH 7.0), and the genome was extracted. Using the extracted M 2014574 genome as a template, primers were designed based on the ppc gene (locus_tag: KPN_04245) on the genome sequence of Klebsiella pneumoniae MGH 78578 registered in NCBI (LOCUS: NC_009648), and the gel was recovered after the PCR reaction. The bands were sequenced, and the target bands were subjected to gene analysis after sequencing, and the similarity with the ppc gene on MGH78578 was 100%. The pck gene (2676bp) of the genome of M 2014574 was knocked out by means of homologous recombination, and the obtained recombinant strain was M 2014574△ppc.
[0029] M 2014574 and M 2014574△ppc were respectively inoculated in 250ml Erlenmeyer flasks at 37°C for anaerobic ...
Embodiment 3
[0033] Example 3. After knocking out the pck gene, the generation of the by-product 2,3-butanediol was significantly reduced and the synthesis of the product 1,3-PD was promoted on the 5L reactor
[0034] The fermentation experiment in a 5L reactor was as follows: the strains (M 2014574△pck, M 2014574) were inserted into a 250ml shake flask (50ml liquid volume) for seed cultivation for 20 hours, and then inserted into a 5L fermenter (2L fermentation liquid volume ), control the fermentation process according to the technological conditions shown below.
[0035] The initial glycerol concentration is 60g / L, the fermentation temperature is 35°C; the ventilation rate is 1.0vvm; the stirring speed is 20rpm; the pH value is controlled by adding NaOH solution to 5.5-7.5 during the fermentation process. In each period of fermentation, the glycerin concentration is controlled at 10-60 g / L by supplementing glycerin solutions with different concentrations, and the fermentation ends in 30...
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