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High proportion psendomonas syringae production strain and its fermentation method to produce psendomonas syringae

A technology of coronatin and bacterial strains, which is applied in the field of microorganisms and their fermentation production compounds, can solve the problems of high cost and unsuitability for industrial production, and achieve the effect of high coronatin production

Inactive Publication Date: 2006-06-14
蔡祝南 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The method of chemical synthesis by Ichihara Dimin et al. (1998) is successful, and it needs about 15 steps of reaction, but the cost is very high, and it is not suitable for industrial production

Method used

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  • High proportion psendomonas syringae production strain and its fermentation method to produce psendomonas syringae
  • High proportion psendomonas syringae production strain and its fermentation method to produce psendomonas syringae
  • High proportion psendomonas syringae production strain and its fermentation method to produce psendomonas syringae

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Take the Y-2018 strain and the starting strain 011 slant strains, inoculate them on the slant medium, and culture them at 26-28°C for 2 days before use. The slant medium is: glucose 1g, beef extract 0.3g, peptone 0.5g, agar 1.5g, water 100mL, pH 7.4, sterilized at 121°C for 30 minutes.

[0046] Make 2 bottles of 100ml seed medium in 500mL Erlenmeyer flasks, one bottle is inserted into the Y-2018 strain, and the other bottle is inserted into the starting strain 011, cultivated on a shaker at 26°C for 24 hours, the speed of the rotary shaker is 160r / min, and the seeds of the shaker flask are long Ready to use later. The seed medium is: 1g of white sugar, 2g of corn steep liquor, 0.1g of ammonium sulfate, 0.1g of yeast extract, 0.15g of dipotassium hydrogen phosphate, 0.08g of magnesium sulfate, 100mL of water, pH7.4, sterilized at 121 for 30 minutes.

[0047] Make two kinds of fermentation medium, one is No. 1 medium without organic nitrogen, the formula is: white sugar ...

Embodiment 2

[0052] The formula of the culture medium used in this test is the same as that of Example 1.

[0053] Inoculate the strains of the Y-2018 strain stored in a refrigerator at 4°C on the slant medium, and cultivate them at 28°C for 48 hours to activate them. Then connect two bacterial loops to 100mL seed medium in 500mL Erlenmeyer flasks, inoculate 3 bottles, and culture on a shaker at 26°C for 24 hours. After the seeds grow well, insert in 500mL Erlenmeyer flask 100mL fermentation medium (No. 2 medium), the inoculum size is 2%, 100 bottles in total, 28 ℃ shaker culture 4 days, merge 100 bottles to get 10L fermented liquid, use The peak area of ​​coronatine was measured by high performance liquid chromatography. As in Example 1, the concentration of coronatin obtained from the standard sample was 36ug / mL, and the total amount of 10L coronatin was 360mg. The fermented liquid is adjusted to pH 3 and filtered, then flows through the macroporous resin (Yangzhou Pharmaceutical Factor...

Embodiment 3

[0055] According to the seed cultivation method of Example 2, obtain 3 bottles of logarithmically grown seeds (100 mL in 500 mL Erlenmeyer flasks), then make 3 bottles of 1000 mL seed culture medium in 3000 mL Erlenmeyer flasks, and insert 100 mL of long-growing seeds respectively, at 26 ° C. Cultivate on a shaker for 18-20 hours before use. Then make the fermentation medium (i.e. No. 2 medium) of the fermenter, and add 0.03% foam enemy, prepare 150L fermentation culture liquid in the 200L fermenter, cool to 28 ℃ through sterilization, insert 3 bottles of 3000mL 1000mL long Good seeds, cultivated at 26°C for 4 days, the ventilation rate is 0.8v.v.m for 0-16 hours, 1.2v.v.m for 17-80 hours, and 0.9v.v.m for 81-96 hours. The mature fermented liquid was determined by high performance liquid chromatography, and the content of coronatin obtained by the same method as in Example 1 was 41.42ug / mL, and the total content of 150L was 6.213g. The fermented liquid is adjusted to pH 3, ad...

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Abstract

The present invention provides a new bacterial strain capable of fermenting and producing high-component coronatin, that is, Pseudomonas syringae (Psendomonas syringae) Y-2018 bacterial strain, and the preservation number is CGMCC NO.1105. The weakness of the wild strain that must be low temperature to accumulate coronatin. The present invention also provides a method for industrialized production of coronatin, which comprises preparing the Pseudomonas syringae Y-2018 strain through slant strains, culturing the seeds in shaking flasks and aerated fermentation to obtain a fermented liquid containing coronatin. Concentrate, extract, crystallize and recrystallize to obtain crystal powder and non-crystallized mother liquor. This method can use organic nitrogen medium for fermentation, and the use of organic nitrogen not only increases the amount of bacteria, but also can synthesize more coronatine, thereby overcoming the defects that wild strains are not resistant to organic nitrogen, and organic nitrogen inhibits the accumulation of coronatin .

Description

technical field [0001] The present invention relates to the field of microorganisms and their fermentative production of compounds. Specifically, it relates to a new bacterial strain for fermentative production of coronatin and a method for fermentative production of coronatin by using the bacterial strain. Background technique [0002] Coronatin is a renamed Chinese name. In the 1990s, Chinese plant pathologists called it coronatoxin. Now we know that coronatin is not only toxic to plants, but also has a very high toxicity to plants. Physiological activity, so it is more appropriate to be called coronatin. The English name is coronatine. In 1977, Ichikara et al. discovered that Pseudomonas syingae pv.alropurea can cause discoloration of Lolium multiflorum, which is caused by a compound produced by the bacteria. Ichihara This compound is named coronatine, because the old name of the bacterium is Ps.corofaciens var.aropurpurea, which is composed of the prefix of its specie...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12P1/04
Inventor 蔡祝南
Owner 蔡祝南
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