Aspergillus niger NY-1 as well as improvement technique for improving insecticidal activity of aspergillus niger NY-1 and application of aspergillus niger NY-1

A technology of NY-1 and Aspergillus niger, applied in the field of chemical epigenetic modification, can solve the problems of delay, difference of metabolic pathways of endophytes, stop of secondary metabolism, etc., and achieve the effect of obvious effect and low cost.

Active Publication Date: 2014-08-20
QINGDAO UNIV OF SCI & TECH
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  • Abstract
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Problems solved by technology

[0002] Symbiotic fungi exhibit abundant secondary metabolism capabilities, and some secondary metabolites also exhibit insecticidal activity. However, when the endophytic bacteria fermentation method is used to produce natural insecticides, on the one hand, after the endophytic bacteria leave the plant, Secondary metabolism is often stopped or delayed; on the other hand, the metabolic pathways of endophytes will be different under autobiotic and symbiotic conditions
When Bok J.W et al. studied the genome of Aspergillus nidulans, they found that there were 1 terpene, 2 indole alkaloids, 14 non-ribosomal polypeptides and 27 polyketides in the genome of the fungus. The gene exists, but when it is fermented and cultivated under experimental conditions, only a few secondary metabolites can be expressed, which seriously hinders the research and development of fungal natural medicines (Chem Biol, 2006, 13(1): 31-37. )

Method used

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  • Aspergillus niger NY-1 as well as improvement technique for improving insecticidal activity of aspergillus niger NY-1 and application of aspergillus niger NY-1

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0012] Example 1: Classification of Aspergillus niger (Aspergillus niger) NY-1 strain derived from coastal plants.

[0013] 1) Bacteria culture

[0014] According to the routine culture method of the microbiology laboratory, a small amount of Aspergillus niger (Aspergillus niger) NY-1 preserved in the glucose-agar medium was picked, inoculated on the surface of the PDA plate, and cultured at 25°C for 4 days. The initial colony was spreading and growing, and the whole was Black, with white mycelium, late spores are black, carpet-like, radial.

[0015] Preliminary identification of strains

[0016] According to the literature method (Journal of Northeast Agricultural University, 2007, 38, (1): 101-106), the ITS segment of 18S-28SrDNA was amplified by PCR, and the sequencing results were compared with the corresponding sequences of known strains in the Genebank database. The similarity of strain Aspergillus niger (Accession number: KF358715.1) was 100%. 序列信息如下:ACTGGGATCTACTGATC...

Embodiment 2

[0017] Example 2: Fermentation of Aspergillus niger NY-1 strain.

[0018] 1) Fermentation culture

[0019] Strain culture: First, according to the routine culture method of the microbiology laboratory, pick a small amount of Aspergillus niger NY-1 (Aspergillus niger) preserved in the glucose-agar medium, inoculate it on the surface of the PDA plate, and cultivate it at 25°C for 4 days as a large-scale fermentation The cultured strains are ready for use.

[0020] Next, cut 4cm from the seed plate 2 Aspergillus niger NY-1 (Aspergillus niger) strains of different sizes were put into sterilized 2000mL Erlenmeyer flasks filled with liquid culture medium, left to stand at room temperature for 22 days, and then sterilized with ethyl acetate for later use.

[0021] The liquid medium is potato 60g / bottle, glucose 6g / bottle, yeast extract 0.9g / bottle, peptone 1.5g / bottle, 5-azacytidine 2.44μg / bottle, distilled water 300mL / bottle.

[0022] 2) Separation and purification of fermentatio...

Embodiment 3

[0024] Example 3: Insecticidal activity test.

[0025] Artemia (brine shrimp), also known as Saltwater Artemia, belongs to Arthropoda, Crustacea, Anocaria, Saltwater Artemia, and Artemia in taxonomy. Xiao Yongtang from Xiamen University used Artemia as the object of toxicology experiments, and studied the anti-insect activity of a total of 188 marine fungal metabolites collected from mangroves in Zhejiang and Fujian Provinces (Journal of Xiamen University, 2005, 44(6): 847- 850); Liu Yong et al. (Acta Ecotoxicology, 2011, 6(5):557-560) determined the safety and ecotoxicological evaluation of 7 pyrethrins on the non-target organism Artemia. Therefore, the experimental method for the acute toxicity of Artemia as a poison is simple, easy to operate, fast and highly feasible. (Plant Medica, 1991, 45:31-34; Pesticides, 2011, 50(4):261-262, 275).

[0026] 1) Hatching of Artemia

[0027] Take a spoonful of about 200mg of Artemia eggs and place them in 500ml of aged seawater for ve...

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Abstract

The invention relates to epigenetic inheritance modification, and in particular relates to an aspergillus niger NY-1 as well as an improvement technique for improving the insecticidal activity of the aspergillus niger NY-1 and application of the aspergillus niger NY-1. The aspergillus niger NY-1 was collected in China Center for Type Culture Collection (CCTCC) on April 28th, 2014 with the collection number CCTCC NO: M2014169. The chemical epigenetic inheritance modification technique is used for improving the insecticidal activity of an NY-1 fermentation product and is applied to a microbial pesticide. The strain is separated from burdock collected in Anqiu, Shandong; an ITS zone is amplified through PCR, and the sequencing result shows that the similarity of the strain is 100% compared with a strain Aspergillus niger (Accession number: KF358715.1) in a Genebank database. An insecticidal silent gene of the aspergillus niger NY-1 is activated by adopting a chemical epigenetic modifier, so that the insecticidal activity of the NY-1 is improved, and the NY-1 is applied to the microbial pesticide. Compared with gene control and other silent gene activation technologies, the method has the advantages of quickness, low cost, obvious effect and the like.

Description

technical field [0001] The invention relates to chemical epigenetic modification technology, more specifically, an Aspergillus niger NY-1 strain and the improvement technology and application of improving its insecticidal activity. After strain NY-1 is treated with chemical epigenetic modifiers, the extract of its fermentation broth has higher insecticidal activity. Background technique [0002] Symbiotic fungi exhibit abundant secondary metabolism capabilities, and some secondary metabolites also exhibit insecticidal activity. However, when the endophytic bacteria fermentation method is used to produce natural insecticides, on the one hand, after the endophytic bacteria leave the plant, Secondary metabolism is often stopped or delayed; on the other hand, endophytes have different metabolic pathways under autobiotic and symbiotic conditions. When Bok J.W et al. studied the genome of Aspergillus nidulans, they found that there were 1 terpene, 2 indole alkaloids, 14 non-ribos...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14A01N63/04A01P7/00C12R1/685
Inventor 李昉
Owner QINGDAO UNIV OF SCI & TECH
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