Avermectin B1a high-yielding strain and application thereof

A technology of avermectin and high-yielding strains, applied in the directions of bacteria, microorganisms, biochemical equipment and methods, etc., can solve the problems of complex mutagenesis process, adverse effects on human health and environmental protection, etc., and achieves good genetic stability and major Effects of social significance and economic value

Inactive Publication Date: 2012-08-15
NANJING UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, because most of the mutagenesis uses highly toxic organic reagents with carcinogenic effects, and the mutagenesis process is complicated, it will have adverse effects on human health and environmental protection. The above technical solutions need to be further improved.

Method used

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  • Avermectin B1a high-yielding strain and application thereof
  • Avermectin B1a high-yielding strain and application thereof
  • Avermectin B1a high-yielding strain and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1: This example illustrates the method of mutagenizing the starting strain of Streptomyces avermitilis.

[0049] The Streptomyces avermitilis AVE07 preserved in the laboratory was used as the starting strain for mutagenesis, and the specific steps were as follows:

[0050] (a) Preparation of monospore suspension: take the fresh slant of Streptomyces avermitilis cultured at 26-30°C for 6-7 days, add 10 mL of sterile water, scrape and wash the spores and pour them into a 250-mL Erlenmeyer flask with certain glass beads for shaking Shake well for 20min (250rpm), break up the spore chains, filter with three layers of degreasing gauze, count the filtrate with a hemocytometer, and adjust the spore concentration by 10 6 individual / ml.

[0051] (b) Low-energy nitrogen ion implantation-lithium chloride compound mutagenesis: Take 0.1 mL of the spore suspension in step (a) and evenly spread it on a sterile empty plate, and perform low-energy nitrogen ion implantation on t...

Embodiment 2

[0060] Example 2: This example illustrates the passage stability of mutant strains.

[0061] In the fermentation medium with glucose as the available carbon source and cornstarch as the late-acting carbon source, the subculture stability of the mutant strain Streptomyces avermitilis AVE07-N2-16515 was detected, and the results of the subculture fermentation test of the strain are shown in Table 1:

[0062] Table 1 Genetic stability test of high-yielding mutant strain Streptomyces avermitilis AVE07-N2-16515

[0063]

[0064] It can be seen from the experimental results that after 5 passages, the avermectin B1a titer of the mutant strain Streptomyces avermitilis AVE07-N2-16515 is relatively stable, and has good passage stability, which can be used as a production product for further research and development. strain.

Embodiment 3

[0065] Example 3: This example illustrates the process of producing abamectin B1a by fermentation of the abamectin B1a high-yielding strain AVE07-N2-16515.

[0066] The medium formula used in this embodiment:

[0067] Plate medium: soluble starch 1.0%, (NH 4 ) 2 SO 4 0.25%, NaCl 0.05%, MgSO 4 ·7H 2 O 0.12%, K 2 HPO 4 ·3H 2 O 0.3%, CaCO 3 0.3%, agar powder 1.5%, pH7.2~7.4, distilled water configuration.

[0068] Slant medium: soluble starch 1.0%, (NH 4 ) 2 SO 4 0.25%, NaCl 0.05%, MgSO 4 ·7H 2 O 0.12%, K 2 HPO 4 ·3H 2 O 0.3%, CaCO 3 0.3%, agar powder 1.5%, pH7.2~7.4, distilled water configuration.

[0069] Seed medium: corn starch 1.5%, glucose 1.0%, peanut cake powder 1.0%, soybean cake powder 0.8%, yeast powder 0.5%, CoCl 2 ·6H 2 O 0.0003%, amylase 4u / g starch, pH 7.0-7.2, tap water configuration.

[0070] Fermentation medium: 6% corn starch, 2% glucose, 1.4% peanut cake powder, 1.2% soybean cake powder, 0.3% yeast powder, CoCl 2 ·6H 2 O 0.0003%, amylase...

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Abstract

The invention discloses an avermectin B1a high-yielding strain, which is classified and named streptomyces avermitilis AVE 07-N2-16515, and is preserved in a China type culture preservation centre (CCTCC) with the preservation number of CCTCC NO: M2012094. The avermectin B1a high-yielding strain disclosed by the invention is obtained by combining low-energy nitrogen ion implantation-lithium chloride (N<+>-LiCl) compound mutation with ultraviolet ray-lithium chloride (UV-LiCl) compound mutation, primarily screening, and performing shake-flask fermentation secondary screening, and is used as a starting strain for next mutation, so that the target strain AVE07-N2-16515 is finally screened. The obtained strain can greatly increase the avermectin B1a component and reduce other components in fermentation products, and is good in hereditary stability. The strain is fermented in a 5L fermentation tank to produce avermectin B1a by utilizing glucose as a quick-acting carbon source and cornstarch as a delayed-action carbon source, the titer can achieve 3048 mu g / m, which is improved by 23.4% as compared with the original starting strain AVE07; and the strain can be applied on industrial production, greatly improves a fermenting unit, and has great economic application value.

Description

technical field [0001] The invention relates to the field of microbial fermentation, in particular to a high-yielding abamectin B1a strain and its mutagenesis method and its application in the fermentative production of avermectin B1a. Background technique [0002] Avermectin (avermectin, AVM) is a kind of 16-membered ring lipid antibiotic produced by liquid fermentation of the actinomycete Streptomyces avermitilis, and it is a new kind of pollution-free biological agricultural antibiotic. It has the characteristics of high efficiency, low toxicity and environmental friendliness. It has good control and killing effects on related pests of vegetables, fruit trees, cotton, rice and other crops. Its insecticidal activity is several to dozens of times higher than that of general chemical pesticides. As an agricultural insecticide, Abamectin has a unique mechanism of action, has contact and stomach poisoning effects on pests, and has a strong penetration effect on target crops, a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P19/62C12R1/465
Inventor 虞龙龚文静崔莉黄思思陈晓双
Owner NANJING UNIV OF TECH
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