KR102189632B1 - Novel fungus Penicillium acidum and use thereof - Google Patents

Abstract

The present invention is a novel fungus Penicillium ashdum ( Penicillium acidum ) and its use, and Penicillium acidum according to the present invention is a strain having an antifungal effect and can achieve biological control by its culture or culture extract, and is an antifungal target, Alternaria Alter. Alternaria alternata ) and Magnaporthe oryzae ) has excellent antifungal effect. Also, Penicillium ashdum acidum ) (Accession No.: KCTC13490BP), its culture medium or its culture filtrate can inhibit or improve specific activity occurring in cells, and has excellent production capacity of specific compounds, making it suitable for compositions in various fields such as pharmacy, food, and beauty. Can be utilized.

Description

Novel fungus Penicillium acidum and use thereof {Novel fungus Penicillium acidum and use thereof}

The present invention relates to a novel Penicillium strain and use thereof.

Various microorganisms such as bacteria, fungi, yeast, and algae exist in nature, and research using them continues. The field of application of microorganisms is very diverse, and representatively, there are therapeutic agents in the medical field, health functional foods in the food field, and cosmetic compositions in the beauty field.

Hundreds of thousands of microorganisms have been discovered and reported to date, but new strains are continuously reported, and discovery of new strains may lead to the invention of new uses using the same. For example, Korean Patent Application Laid-Open No. 10-2016-0087438 discloses a method of preparing fermented products such as paste using a novel penicillium strain.

Korean Patent Application Publication No. 10-2016-0087438

The present invention provides a novel fungus Penicillium acidum , including microbial preparations for antibacterial using the same, pharmaceutical compositions, health functional foods, cosmetic compositions, feed, various types of antibacterial agents, etc. Provides an application plan.

The present invention provides Penicillium acidum (Accession No.: KCTC13490BP).

The pharmaceutical composition of the present invention is Penicillium ash. acidum ) (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) culture solution and penicillium acidum ( Penicillium acidum ) (Accession No.: KCTC13490BP) in the group consisting of the culture filtrate It contains at least one selected as an active ingredient.

The health functional food of the present invention is Penicillium ash acidum ) (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) culture solution and penicillium acidum ( Penicillium acidum ) (Accession No.: KCTC13490BP) in the group consisting of the culture filtrate It contains one or more selected as active ingredients.

The cosmetic composition of the present invention is Penicillium ash acidum ) (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) culture solution and penicillium acidum ( Penicillium acidum ) (Accession No.: KCTC13490BP) in the group consisting of the culture filtrate It contains at least one selected as an active ingredient.

The antifungal microbial preparation of the present invention is a culture medium of Penicillium acidum (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) and Penicillium acidum ( Penicillium acidum). ) (Accession No.: KCTC13490BP) contains one or more selected from the group consisting of the culture filtrate as an active ingredient.

In the microbial preparation for antifungal according to an embodiment of the present invention, the antifungal target of the microbial preparation is Alternaria Alternata ( Alternaria alternata ), Alternaria mali , Aspergillus oryzae ), Botrytis cinerea , Colletotrichum 　 coccodes ), Colletotrichum Groeosporioides gloeosporioides ), Colletotrichum orbiculare ), Cylindrocarpon destructans , Fusarium oxisporum f. sp. Lycopersici ( Fusariumoxyporum f. sp. lycopersici ), Magnaporthe oryzae , Rhizoctonia solani ( Rhizoctonia) 　 solani ), Rhizopus storonifer var. Rhizopus stolonifer var. stolonifer ) may be one or more pathogens selected from the group consisting of, preferably Alternaria ( Alternaria alternata ) or Magnaporthe oryzae , but is not limited thereto.

Penicillium ash according to the present invention acidum ) (Accession No.: KCTC13490BP) is a strain having an antifungal effect and can achieve biological control by its culture or culture extract, and the antifungal targets of Alternaria alternata and Magnaporthe oryzae ) has excellent antifungal effect.

Also, Penicillium ashdum acidum ) (Accession No.: KCTC13490BP), its culture medium or its culture filtrate can inhibit or improve specific activity occurring in cells, and has excellent production capacity of specific compounds, so it is suitable for compositions in various fields such as pharmaceutical, food, and beauty. Can be utilized.

1 is a novel fungus Penicillium ashdum ( Penicillium acidum ) Shows the phylogenetic tree of CNUFCDLW4-1 and CNUFC-DLW4-2 strains.
Figure 2 is a novel fungus Penicillium ashdum ( Penicillium acidum ) shows the morphological structure of the CNUFC-DLW4-1 strain [a and e are colonies in Czapek yeast auto lysate agar (CYA) medium, b and f are colonies in malt extract agar (MEA) medium, c and g Is yeast extract sucrose agar (YES) colonies, d and h are creatine sucrose agar (CREA) colonies, i~k and m~r are conidiophores, l and s are conidia Show The scale bar is 20 μm for m, 10 μm for i-l and p, 5 μm for n, o, q and r, and 2 μm for s].
3 is a BenA gene sequence of Penicillium acidum (Accession No.: KCTC13490BP).
Figure 4 is a CaM gene sequence of Penicillium acidum (Accession No.: KCTC13490BP).
5 is a sequence of RPB2 gene of Penicillium acidum (Accession No.: KCTC13490BP).
Figure 6 is Penicillium ashdum ( Penicillium acidum ) relates to the antifungal activity of the CNUFCDLW4-1 strain. A. Magnaporthe oryzae ) (control) for Penicillium ashdum ( Penicillium acidum ) Front view of CNUFCDLW4-1, B. Magnaporthe oryzae ) (Control) of Penicillium acidum CNUFCDLW4-1, C. Alternaria Alternaria alternata ) Penicillium ash against EML-BLDF1-4 (control) acidum ) Front view of CNUFCDLW4-1, D. Alternaria Alternaria alternata ) Back view of Penicillium acidum CNUFCDLW4-1 against EML-BLDF1-4 (control)
Figure 7 is Penicillium ashdum ( Penicillium acidum ) shows the enzymatic activity of CNUFCDLW4-1.
A. Front view of clear zone around colonies formed by culturing Penicillium ash CNUFCDLW4-1 on skim milk agar, B: Clear zone around colonies formed by culturing Penicillium ash CNUFCDLW4-1 on skim milk agar The back side of it.

Hereinafter, with reference to the accompanying table or drawings will be described in detail the penicillium ash ( Penicillium acidum ) strain of the present invention and the antifungal microbial preparation comprising the same.

When the drawings are described, this is provided as an example in order to sufficiently convey the spirit of the present invention to those skilled in the art. Accordingly, the present invention is not limited to the drawings to be presented and may be embodied in other forms, and the drawings may be exaggerated to clarify the spirit of the present invention.

In this case, unless there are other definitions in the technical terms and scientific terms used, they have the meanings commonly understood by those of ordinary skill in the art to which this invention belongs, and the gist of the present invention in the following description and accompanying drawings Descriptions of known functions and configurations that may be unnecessarily obscure will be omitted.

In addition, the singular form used in the specification of the present invention may be intended to include the plural form unless otherwise indicated in the context.

In addition, units used in the specification of the present invention are based on weight, and as an example, the unit of% or ratio means weight% or weight ratio.

In addition, in the specification of the present invention, the expression "includes" is an open-type substrate having a meaning equivalent to expressions such as "have", "include", "have" or "feature", and are further listed. It does not exclude elements, materials or processes that do not exist. Also, “Practically… The expression “consist of” means that the specified element, material or process does not have an unacceptably significant effect on at least one basic and novel technical idea of the invention by another element, material or process not listed. It means something that can exist as a quantity. Also, the expression “consisting of” means that only the described elements, materials or processes are present.

In the present invention, “sample” or “sample” refers to an object for analysis, and has the same meaning throughout the specification.

Hereinafter, the present invention will be described in detail.

The present invention provides a novel fungus Penicillium acidum (Accession No.: KCTC13490BP).

The penny room Solarium ash Doom (Penicillium acidum) is separated from the water collected in a small pond and identification and Solarium ash chamber penny Doom (Penicillium acidum ) sp. nov. It was named CNUFC-DLW4-2, and as of March 05, 2018, it was deposited with the Korea Institute of Biological Resources under the deposit number KCTC13490BP.

Penicillium acidum , a novel fungus of the present invention, is a new strain of Penicillium John Crugi . johnkrugii), Penny room Solarium words value (Penicillium mallochii) and Penny's room Solarium exciter means (P. exsudans) It is highly correlated with HMAS248735, but shows slow growth in certain media, CYA and YES, and produces strong acids in CREA media, which distinguishes them from these.

Penicillium acidum (Accession No.: KCTC13490BP) of the present invention has various biological activity effects, and can produce a large amount of specific substances.

The pharmaceutical composition of the present invention is Penicillium ash. acidum ) (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) culture solution and penicillium acidum ( Penicillium acidum ) (Accession No.: KCTC13490BP) in the group consisting of the culture filtrate It contains at least one selected as an active ingredient.

The health functional food of the present invention is Penicillium ash acidum ) (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) culture solution and penicillium acidum ( Penicillium acidum ) (Accession No.: KCTC13490BP) in the group consisting of the culture filtrate It contains at least one selected as an active ingredient.

The cosmetic composition of the present invention is Penicillium ash acidum ) (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) culture solution and penicillium acidum ( Penicillium acidum ) (Accession No.: KCTC13490BP) in the group consisting of the culture filtrate It contains at least one selected as an active ingredient.

According to an embodiment of the present invention, Penicillium acidum (Accession No.: KCTC13490BP) is a culture medium of Penicillium acidum (Accession No.: KCTC13490BP) as well as the cells themselves, penicillium A culture filtrate obtained by filtering the culture solution of Ashidoum ( Penicillium acidum ) (Accession No.: KCTC13490BP) may also have various biological activity effects.

In addition to the culture solution and the filtrate of the culture solution, a freeze-dried powder of the culture solution, a freeze-dried powder of the culture filtrate, or a solution obtained by dissolving the freeze-dried powder can be used in various fields.

In the present invention, the culture medium may include a substance produced by the cells by culturing the cells in a medium, the cells themselves, and a medium.

In the present invention, the culture filtrate may include a supernatant obtained by centrifuging the culture solution, and a liquid obtained by filtering the supernatant.

Various compositions of the present invention, methods for preparing foods, etc. include the steps of culturing Penicillium acidum (accession number: KCTC13490BP) in a culture medium and Penicillium acidum (Accession number: KCTC13490BP) It may include the step of obtaining and formulating a culture solution obtained by culturing.

The method of preparing the composition of the present invention may further include the step of preparing a supernatant by centrifuging the culture solution to obtain a supernatant of the culture solution in which layer separation has occurred.

The method for preparing the composition of the present invention may further include formulating a culture filtrate obtained by centrifuging the culture solution to filter the supernatant of the culture solution in which layer separation has occurred.

In the present invention, the medium for culturing Penicillium acidum (accession number: KCTC13490BP) is MEA (Blakeslee's malt extract agar) medium, YES (yeast extract sucroseagar) medium, CREA (Czapek yeast autolysateagar) medium and PDA (potato dextrose agar) may be a medium selected from a medium, but is not limited thereto. According to an embodiment of the present invention, it is preferably MEA and CREA medium, and growth may be slow in CYA and YES medium.

The antifungal microbial preparation of the present invention is a culture medium of Penicillium acidum (Accession No.: KCTC13490BP), Penicillium acidum (Accession No.: KCTC13490BP) and Penicillium acidum ( Penicillium acidum). ) (Accession No.: KCTC13490BP) contains one or more selected from the group consisting of the culture filtrate as an active ingredient.

In the microbial preparation for antifungal according to an embodiment of the present invention, the antifungal target of the microbial preparation is Alternaria Alternata ( Alternaria alternata ), Alternaria mali , Aspergillus oryzae ), Botrytis cinerea , Colletotrichum 　 coccodes ), Colletotrichum Groeosporioides gloeosporioides ), Colletotrichum orbiculare ), Cylindrocarpon destructans , Fusarium oxisporum f. sp. Lycopersici ( Fusariumoxyporum f. sp. lycopersici ), Magnaporthe oryzae , Rhizoctonia solani ( Rhizoctonia) 　 solani ), Rhizopus storonifer var. Rhizopus stolonifer var. stolonifer ) may be one or more pathogens selected from the group consisting of, preferably Alternaria alternata or Magnaporthe oryzae , but is not limited thereto.

Plants caused by the fungus exhibit symptoms such as black mold rot, leaf spots, dry spots, ulcers, dry branches, and root rot, and leaves or stems wither, and the quality of fruits deteriorates.

The type of plant fungal diseases caused by the fungus is not particularly limited in the present invention, but may include fungal diseases that can be invented by fungi that can be controlled through the microbial preparation, As a specific example, 　 plant diseases caused by molds such as powdery mildew, gray mold or leaf mold may be mentioned, but are not limited thereto.

In addition, Penicillium acidum (Accession No.: KCTC13490BP) of the present invention can be prepared in a form used in various fields such as pharmaceutical compositions, health functional foods, cosmetics, feed, antibiotics, and the like.

For example, when the composition according to the present invention is prepared in the form of a health functional food or a food additive, the form or type is not particularly limited, and may be prepared by mixing with other ingredients capable of increasing the activity. The type of food is also not particularly limited, and for example, it may include drinks, vitamins, various beverages, processed meats, processed sausages, processed noodles, processed fats and oils, sugars, and foods that are commonly manufactured and sold.

Preferably, it is good to prepare a drink or other beverage-type composition, and various flavoring agents such as sweeteners may be included as additional ingredients to help intake. It may contain carbohydrates having various flavors such as simple sugar, fructose, and glucose, and sugar alcohols such as sorbitol or xylitol may also be included. As an example of the sweetener, not only natural sweeteners but also synthetic sweeteners such as saccharin may be used.

In addition, when the composition according to the present invention is prepared in the form of a pharmaceutical composition or a drug, the form or type is not particularly limited, and may be prepared in various formulations according to an administration or intake method. In addition, excipients, carriers, or diluents commonly used in the relevant field may be appropriately further included when preparing a pharmaceutical composition. Examples of the dosage form may include various dosage forms such as external preparations or injection preparations in addition to oral forms such as tablets, syrup, capsules, and powders. Examples of carriers and excipients include sorbitol, sugar alcohols such as xylitol or malthiol, monosaccharides, polysaccharides, gelatin, water, alcohols, starches, edible rubbers, mineral oils, and various compounds, and mixtures thereof. The types of wetting agents, surfactants, excipients, fillers, binders, etc. generally used in formulation are not particularly limited.

Hereinafter, the content of the present invention will be described in more detail through examples. The examples are only for describing the present invention in more detail, and the scope of the present invention is not limited thereto.

[ Example One]

Identification of fungi

Example 1-1. Sampling and isolation for the identification of new fungi

Freshwater samples were collected from a small pond at Chonnam National University (Gwangju, Korea). The collected sample was transferred to a sterilized 50 ml conical tube and stored at 4°C. Mold bacteria were isolated by serial dilution plating method. Specifically, 1 ml of water, which is a sample, was mixed with 9 ml of sterile distilled water, shaken for 15 minutes at room temperature, and then diluted stepwise to a concentration of 10 ^-1 to 10 ^-4 . 0.1 ml of each dilution was transferred to potato glucose agar medium (PDA medium) (39g potato dextrose agar [Becton, Dickinson, and Co., Sparks, MD, USA]) and incubated at 25° C. for 3 to 7 days. To isolate pure cultures, individual colonies of various types were transferred to PDA plates. The pure strain was stored in a PDA slant tube and 20% glycerol at -80°C.

Example 1-2. DNA extraction, PCR And sequencing

Each of the isolated bacteria was incubated for 3 to 5 days at 25°C in PDA covered with cellophane. Total genomic DNA was extracted using a Solgent Genomic DNA prep kit (Solgent Co. Ltd., Daejeon, Korea). Encoding beta tubulin using primer pair Bt2a/Bt2b (Glass & Donaldson 1995), CMD5/CMD6 (Samson et al.), RPB2-5F / RPB2-7cR (Liu et al., 1999) The BenA partial gene, CaM, was used by amplifying and confirming the CaM partial gene encoding calmodulin and the RPB2 partial gene encoding RNA polymerase II.

Each PCR amplification mixture (total volume, 20 µl) contained 2 µl of strain DNA template (10 ng), 1.5 µl of each primer (5 pM/µl); Taq DNA polymerase, dNTP, buffer, and Accupower® PCR premix (Bioneer Corp., Daejeon, Korea) containing 1 µl and 14 µl of sterile water containing a tracking dye. The PCR product was purified with Accuprep® PCR Purification Kit (Bioneer) according to the manufacturer's instructions. DNA sequencing was performed using an ABI 3700 automated DNA sequencer (Applied Biosystems Inc., Foster City, CA, USA).

[Table 1]

Example 1-3. Phylogenetic analysis

Clustal_X v.1.83 (Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Research: 4876-82) and Bioedit v. 5.0.9.1 Software (Hall TA. BioEdit: a user friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp Ser 1999;41:95-8). Phylogenetic analysis was performed using MEGA 6 software (Tamura K, Stecher G, Peterson D, Filipski A, KumarS. 2013. MEGA6: molecular evolutionary genetics analysis version 6.0. Molecular Biology and Evolution 30: 2725-2729).

The number of phylogenies, including CNUFC-DLW4-1, CNUFC-DLW4-2, and the section Sclerotiora related species, was determined by the maximum likelihood analysis for BenA, CaM and RPB2. It was constructed using, and Penicillium levitum was selected as the out group.

Sequence matching rates were obtained by searching the Basic Local Alignment Search Tool (BLASTn) (NCBI) for each strain. The reliability of the inner branch was evaluated using a p-distance substitution model using 1000 bootstrap replicates.

Example 1-4. Morphological analysis

Blakeslee's malt extract agar (MEA; Blakeslee, 1915), yeast extract sucrose agar (YES; Frisvad, 1981), Czapek yeast auto lysate agar ( CYA; Pitt, 1979) and creatine sucrose agar (CREA; Frisvad 1981). Plates (3 replicates for each medium) were inoculated at three-points. After culturing for 7 days, growth was confirmed in the dark at 5°C, 20°C, 25°C, 30°C, and 37°C using Rayner's color charts (the color charts of Rayner (1970)). The sample was mounted on a lactophenol solution (Junsei Chemical Co. Ltd., Tokyo, Japan) and observed with an Olympus BX51 microscope equipped with a DIC optical device (Olympus, Tokyo, Japan).

The fine homogeneous structure was observed with a scanning electron microscope (SEM) (Hitachi S4700; Hitachi, Tokyo, Japan). The isolated strain was fixed in 2.5% paraformaldehyde-glutaraldehyde (paraformaldehyde-glutaraldehyde) in 0.05M phosphate buffer (pH 7.2) for 2 hours, and then washed with cacodylate buffer (Junsei Chemical Co. Ltd.) I did. The cell membrane was preserved by fixing the sample in 1% osmium tetroxide diluted in cacodyl hydrochloric acid for 1 hour (Electron Microscopy Sciences, Hatfield, PA, USA). The sample was washed again with cacodyl hydrochloric acid buffer, dehydrated with graded ethanol (Emsure, Darmstadt, Germany) and isoamyl acetate (Junsei Chemical Co. Ltd.), and dried in a fume hood. Made it.

Finally, the samples were sputter coated with gold and observed using a Hitachi S4700 field emission scanning electron microscope (FE-SEM) at the Korea Basic Science Research Institute.

Example 1-5. Phylogenetic analysis results

In order to analyze the phylogenetic relationship of the related species of the new strain, according to the sequence analysis comparing the ITS rDNA sequence with the related species extracted from GeneBank, Penicillium acidum CNUFC-DLW4-1 and CNUFC-DLW4- 2 was deposited in the NCBI database, and analyzed using it. Analysis was performed using ITS, BenA (SEQ ID NO: 9), CaM (SEQ ID NO: 10), and RPB2 (SEQ ID NO: 11) genes ([Table 2]). BenA is a gene that encodes beta tubulin, CaM is a gene that encodes calmodulin, and RPB2 is a gene that encodes RNA Polymerase II.

When comparing the ITS rDNA sequences of the CNUFC-DLW4-1 and CNUFC-DLW4-2 strains with related species extracted from GenBank, sequence analysis by BLASTn search showed that CNUFC-DLW4-1 and CNUFC-DLW4-2 were Penicillium sp. .( Penicillium sp.) YZ02 (GenBank accession number JQ809682) and Penicillium sp. sp.) showed the closest correlation with PE125 (GenBank accession number JX875942).

Penicillium John Crugi johnkrugii ) DAOM 239945 and Penicillium johnkrugii The BenA sequence of DAOM 239942 showed 97.2% (412/424 bp) sequence homology with the isolate's BenA. Penny room Solarium words value (Penicillium mallochii) DAOM 239919 and Penny room Solarium words value (Penicillium mallochii ) The CaM sequence of DAOM 239917 showed 97.3% (442/454 bp) sequence homology with the CaM of the isolate. Penicillium Exsudans ( P. exsudans ) The RPB2 sequence of HMAS248735 showed a sequence homology of 99.1% (918/926 bp) with RPB2 of the isolate.

Phylogenetic trees were deduced from the data set for BenA, CaM and RPB2 of CNUFC-DLW4-1 and CNUFC-DLW4-2 and related species belonging to the Sclerotiora section extracted from GenBank (Figure 1). Phylogenetic analysis placed the isolated fungi in the Sclerotiora section, and the isolated fungi formed a separate clade, showing that the isolated fungi were separate Penicillium species.

Example 1-6. Morphological analysis results

Fungal colonies grew rapidly in MEA and reached a diameter of 31 to 32 mm in 7 days at 25°C. Conidiophores are formed as a single monoverticillate, and the stipes are gentle, vary in length, form partitions, have a width of 2.5 to 4.0 µm, and the spore forming organ, phialide, is Smooth, flask-shaped, 5-12 per verticil, conidia slightly coarse, spherical to hemispherical, and the size is 2.5-3.5㎛ Х 2.0-3.0㎛ (Fig. 2).

The new strain showed slow growth when cultured in CYA and YES media, and sclerotia were formed in some media, resulting in Penicillium johnkrugii . And Penicillium mallochii . Penicillium johnkrugii And Penicillium mallochii did not produce a strong acid, whereas the new strain produced a strong acid in CREA medium.

The new fungus Penicillium acidum ) CNUFC-DLW4-1 and CNUFC-DLW4-2, of which CNUFC-DLW4-2 was deposited with the Korea Research Institute of Bioscience and Biotechnology Biological Resource Center (KCTC).

Deposit number: KCTC13490BP

Deposit date: 2018.03.05.

Depositary: Korea Research Institute of Bioscience and Biotechnology Biological Resource Center

[ Example 2]

new Penicillium Ashdum( Penicillium acidum )of Active check

Example 2-1. Penicillium Ashdum ( Penicillium acidum ) CNUFC - DLW4 Measurement of antifungal activity of -2 ( antifungal activity assay)

Penicillium ashdum acidum ) The antifungal activity of CNUFC-DLW4-2 was measured in PDA medium using a double culture method.

Alternaria Alternaria , a fungal pathogen alternata ) EML-BLDF1-4 and Magnaporthe oryzae ( Magnaporthe oryzae ) of the mycelial plug (mycelial plug) is placed in the center of each PDA plate, Penicillium ashdum CNUFC-DLW4-2 strain culture solution from the pathogenic fungi 2.5 It was placed at a distance of cm.

All plates were incubated for 5-7 days in a dark room at 25°C, and each experiment was repeated three times. The control plate placed only the hyphae plugs of each fungal pathogen.

Thereafter, the width of the inhibitory region of the fungal pathogen and Penicillium ashdum CNUFC-DLW4-2 was measured in millimeters and used as antifungal activity.

The inhibition rate (%) is calculated as follows.

Inhibition rate (%) = (R-r)/R × 100

R is for mycelial growth in the opposite direction of CNUFC-DLW4-1 culture

R is the mycelial growth in the direction of CNUFC-DLW4-1 culture.

As a result, the culture medium of the Penicillium ashdum CNUFC-DLW4-2 strain was used as a fungal pathogen Alternaria alternata in PDA medium. alternata ) It was confirmed that the hyphae growth of EML-BLDF1-4 and Magnaporthe oryzae was significantly inhibited by 30.4% and 42.5%, respectively, showing high antifungal activity (Table 2, FIG. 6).

As a result, the culture medium of the Penicillium ashdum CNUFC-DLW4-2 strain was used as a fungal pathogen Alternaria alternata in PDA medium. alternata ) It was confirmed that the growth of EML-BLDF1-4 and Magnaporthe oryzae was significantly suppressed.

[Table 2]

Example 2-2. Penicillium Ashdum ( Penicillium acidum ) CNUFC - DLW4 Measurement of proteolytic activity of -2 ( proteolytic activity assay)

The agar plate containing 2% agar was inoculated with the strain of Penicillium ashdum CNUFC-DLW4-2 of the present invention and supplemented with 2% skim milk. The plate was incubated for 4 days (3-5 days) at 25°C.

The screening results for the proteolytic activity of the novel Penicillium ashdum CNUFC-DLW4-2 are shown in FIG. 6. From this, the penicillium ash of the present invention produced a proteolytic enzyme after 24 hours incubation, which could be confirmed from a clear zone surrounding the colony.

From the above description, those skilled in the art to which the present invention pertains will be able to understand that the present invention can be implemented in other specific forms without changing the technical spirit or essential features thereof. In this regard, the embodiments described above are illustrative in all respects and should be understood as non-limiting. The scope of the present invention should be construed that all changes or modifications derived from the meaning and scope of the claims to be described later rather than the above detailed description and equivalent concepts are included in the scope of the present invention.

Korea Research Institute of Bioscience and Biotechnology KCTC13490 20180305

<110> CHONNAM NATIONAL UNIVERSITY <120> Novel fungus Penicillium acidum and use thereof <130> P-0305-KCTC13490BP <160> 11 <170> KoPatentIn 3.0 <210> 1 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> ITS rDNA forward primer (ITS-1) <400> 1 tccgtaggtg aacctgcgg 19 <210> 2 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> ITS rDNA reverse primer (ITS-4) <400> 2 tcctccgctt attgatatgc 20 <210> 3 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> BenA Forward primer (Bt2a) <400> 3 ggtaaccaaa tcggtgctgc tttc 24 <210> 4 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> BenA Reverse primer (Bt2b) <400> 4 accctcagtg tagtgaccct tggc 24 <210> 5 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> CaM Forward primer (CMD5) <400> 5 ccgagtacaa ggargccttc 20 <210> 6 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> CaM Reverse primer (CMD6) <400> 6 ccgatrgagg tcatracgtg g 21 <210> 7 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> RPB2 Forward primer (5F) <400> 7 gaygaymgwg atcayttygg 20 <210> 8 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> RPB2 Reverse primer (7CR) <400> 8 cccatrgctt gyttrcccat 20 <210> 9 <211> 423 <212> DNA <213> Artificial Sequence <220> <223> Penicillium acidum sp. nov. CNUFC-DLW4-2 (BenA gene [encoding) beta tubulin]) <400> 9 tcttcaattg atgaaaccag gaacctttga ctgacttgtt atataggcag aacattgcta 60 gcgagcatgg cctcgacggc gagggccagt aagtatcaat ttgcgtggaa ttgggtgata 120 tgagaatggc ggtctgatat ttttctttag cttcactggc cagtccgacc tccagctcga 180 gcgcatgaac gtctacttca accacgtaag tgtggaatca acactcgata cccgtcaacc 240 tcatctaata taatggtttt ccataggcct ctggtgaccg ttacgttccc cgtgccgtcc 300 tggtcgactt ggagcccggt accatggacg ctgtccgtgc cggtcctttc ggcaagcttt 360 tccgccccga caacttcgtc ttcggtcagt ccggtgctgg taacaactgg gccaagggtc 420 act 423 <210> 10 <211> 432 <212> DNA <213> Artificial Sequence <220> <223> Penicillium acidum sp. nov. CNUFC-DLW4-2 (CaM gene [encoding) calmodulin]) <400> 10 ccggtaatag gacaaggatg gcgatggtta gtgcgaccgt cggcgatttt ttaaattcca 60 taccgactgg agtaaaacca ctgagagaac cgaataactg agaccgatcg atctatagga 120 caaatcacca ccaaggagct gggcactgtc atgcgctccc tcggccagaa cccctccgag 180 tccgagcttc aggacatgat caacgaggtc gatgccgaca acaacggcac cattgacttc 240 cctggtacga tttttcttcc gatcgaatga ccccgtgctc cctccggata gatgttaacg 300 tgcggcacag agttcctgac catgatggcc cgtaagatga aggacaccga ctccgaggag 360 gagatccgtg aggccttcaa ggttttcgac cgcgacaaca acggtttcat ttccgctgcc 420 gagctgcgcc ac 432 <210> 11 <211> 934 <212> DNA <213> Artificial Sequence <220> <223> Penicillium acidum sp. nov. CNUFC-DLW4-2 (RPB2 gene [encoding RNA) polymerase II]) <400> 11 ttttccgtgt tcttttcacc cgtgtcaccc gtgatctcca gcgttacgtg cagcgctgcg 60 ttgagactgg ccgcgaaatt taccttaacg tgggtcttaa ggctgctaca ctcactggtg 120 gtctcaagta cgccctggct actggtaact ggggagagca gaagaaggct gccagcgcaa 180 aggccggtgt gtctcaagtg ctgagtcgtt acacatatgc ttctacctta tcccatcttc 240 ggcgtaccaa cactcctatc ggacgtgatg gtaagattgc caaacctcgc caacttcaca 300 atactcactg gggtctggtt tgcccggccg aaacacctga aggacaggct tgtggtctgg 360 tcaagaactt ggcacttatg tgctatatca cagtcggaac acctagtgag cccattatcg 420 atttcatgat tcagcgtaac atggaggtcc ttgaagagtt cgaaccgcag gtgacaccaa 480 acgctaccaa ggtgttcgtg aacggtgtgt gggttggtat ccatcgcgat ccctcccacc 540 tggtcaacac tatgcagaac ctccgtcgcc gcaacatgat ttcgaacgaa gtcagtttga 600 ttcgtgacat tcgtgagcga gagttcaaga tctttactga tgctggtcgt gtttgccgtc 660 ctctcttcgt tgttgacaat gaccccaaga gcgaaaatgc gggatctctg gttctcaaca 720 aggaacacat tcgcaagttg gagcaggaca aggacctgcc aaccgacatg gacttggagg 780 aacgccggga gcgctacttc ggatgggagg gtctggttcg atccggtgcc gttgaaattg 840 tggatgcaga ggaggaagag accatcatga tcgtcatgac acccgaagat ctagagatct 900 ccaagcaact gcaagccggc tacaccctgc caac 934

Claims (6)

Penicillium acidum CNUFC-DLW4-2 (accession number: KCTC13490BP). Penicillium acidum CNUFC-DLW4-2 (Accession No.: KCTC13490BP), Penicillium acidum CNUFC-DLW4-2 (Accession No.: KCTC13490BP) culture and penicillium ash doom ( Penicillium acidum ) CNUFC-DLW4-2 (Accession No.: KCTC13490BP) a composition for protein degradation comprising one or more selected from the group consisting of culture filtrate as an active ingredient. delete delete Penicillium acidum CNUFC-DLW4-2 (Accession No.: KCTC13490BP), Penicillium acidum CNUFC-DLW4-2 (Accession No.: KCTC13490BP) culture and penicillium ash doom ( Penicillium acidum ) CNUFC-DLW4-2 (accession number: KCTC13490BP) as an antifungal microbial preparation containing at least one selected from the group consisting of filtrate culture,
The antifungal target is Alternaria alternata ( Alternaria alternata ), Magnaporthe oryzae ( Magnaporthe oryzae ) or Colletotricum Groeosporioides ( Coletotrichum gloeosporioides ) is a microbial preparation for antifungal. delete

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