WO2022088205A1 - 一种木霉菌扩繁生产菌肥的方法 - Google Patents
一种木霉菌扩繁生产菌肥的方法 Download PDFInfo
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- trichoderma
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- bacterial fertilizer
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- 241000223259 Trichoderma Species 0.000 title claims abstract description 40
- 230000001580 bacterial effect Effects 0.000 title claims abstract description 20
- 239000003337 fertilizer Substances 0.000 title claims abstract description 18
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 12
- 238000000034 method Methods 0.000 claims abstract description 18
- 239000000203 mixture Substances 0.000 claims abstract description 18
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 16
- 239000000725 suspension Substances 0.000 claims abstract description 15
- 238000003756 stirring Methods 0.000 claims abstract description 13
- 239000008223 sterile water Substances 0.000 claims abstract description 10
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 claims abstract description 8
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 claims abstract description 7
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims abstract description 7
- 235000005822 corn Nutrition 0.000 claims abstract description 7
- 238000000855 fermentation Methods 0.000 claims abstract description 7
- 230000004151 fermentation Effects 0.000 claims abstract description 7
- 239000007788 liquid Substances 0.000 claims abstract description 7
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims abstract description 4
- 229930006000 Sucrose Natural products 0.000 claims abstract description 4
- 229910000019 calcium carbonate Inorganic materials 0.000 claims abstract description 4
- 229940116007 ferrous phosphate Drugs 0.000 claims abstract description 4
- 235000013312 flour Nutrition 0.000 claims abstract description 4
- 229910000155 iron(II) phosphate Inorganic materials 0.000 claims abstract description 4
- SDEKDNPYZOERBP-UHFFFAOYSA-H iron(ii) phosphate Chemical compound [Fe+2].[Fe+2].[Fe+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O SDEKDNPYZOERBP-UHFFFAOYSA-H 0.000 claims abstract description 4
- 239000005720 sucrose Substances 0.000 claims abstract description 4
- 235000015099 wheat brans Nutrition 0.000 claims abstract description 4
- ODINCKMPIJJUCX-UHFFFAOYSA-N Calcium oxide Chemical compound [Ca]=O ODINCKMPIJJUCX-UHFFFAOYSA-N 0.000 claims description 20
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 20
- 239000000463 material Substances 0.000 claims description 15
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims description 10
- 239000000292 calcium oxide Substances 0.000 claims description 10
- 235000012255 calcium oxide Nutrition 0.000 claims description 10
- 239000004202 carbamide Substances 0.000 claims description 10
- 235000013877 carbamide Nutrition 0.000 claims description 10
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 10
- 235000019341 magnesium sulphate Nutrition 0.000 claims description 10
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 10
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 10
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 10
- 238000010790 dilution Methods 0.000 claims description 7
- 239000012895 dilution Substances 0.000 claims description 7
- 238000005406 washing Methods 0.000 claims description 6
- 241000894006 Bacteria Species 0.000 claims description 5
- 241000209140 Triticum Species 0.000 claims description 3
- 235000021307 Triticum Nutrition 0.000 claims description 3
- 238000007605 air drying Methods 0.000 claims description 3
- 230000001954 sterilising effect Effects 0.000 claims description 3
- 238000004659 sterilization and disinfection Methods 0.000 claims description 3
- 241000209149 Zea Species 0.000 claims 2
- 239000006228 supernatant Substances 0.000 claims 1
- 239000013067 intermediate product Substances 0.000 abstract description 6
- 240000008042 Zea mays Species 0.000 abstract description 5
- 230000008635 plant growth Effects 0.000 abstract description 4
- 239000002994 raw material Substances 0.000 abstract description 3
- 230000001902 propagating effect Effects 0.000 abstract 2
- 239000001963 growth medium Substances 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 7
- 239000000047 product Substances 0.000 description 5
- 238000000464 low-speed centrifugation Methods 0.000 description 2
- 239000000575 pesticide Substances 0.000 description 2
- 239000002689 soil Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 230000003042 antagnostic effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 230000003032 phytopathogenic effect Effects 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
Definitions
- the invention relates to the technical field of biological bacteria production, in particular to a method for the propagation of Trichoderma to produce bacterial fertilizer.
- Trichoderma belongs to the fungal phylum, Deuteromycota, Hychospora, Hychospora, Trichoderma, and is widely present in soil under different environmental conditions. Since the mid-19th century, humans have had a preliminary understanding of Trichoderma, but it was not until the 1960s that the taxonomic status of Trichoderma was determined. Most Trichoderma can produce a variety of bioactive substances that have antagonistic effects on phytopathogenic fungi, bacteria and insects, such as cell wall degrading enzymes and secondary metabolites, and can improve the stress resistance of crops, promote plant growth and improve agricultural products. Therefore, it is widely used in biological control, biological fertilizers and soil conditioners. Due to the serious negative impact of chemical pesticides on the environment, the environmentally friendly biological pesticide Trichoderma has received extensive attention.
- the purpose of the present invention is to provide a kind of method for Trichoderma to multiply and produce bacterial fertilizer, to solve the problem raised in the above-mentioned background technology.
- a kind of method for Trichoderma propagation to produce bacterial fertilizer may further comprise the steps:
- Step 1 inoculate the Trichoderma strain into the PDA medium and cultivate for 3-6 days to obtain the Trichoderma species;
- Step 2 Take the weight fractions of 10 parts of corn flour, 10 parts of wheat bran, 5 parts of bran, 2 parts of sucrose, 1 part of calcium carbonate and 0.1 part of ferrous phosphate, add sterile water, and add them together in a mixer to fully stir and mix to prepare the mixture. into a liquid fermentation broth, adjust the pH to 5.5-6, and then squeeze out excess water, under high pressure and high temperature environment, carry out sterilization treatment for 30-40min, and naturally cool to obtain culture material;
- Step 3 Add the Trichoderma species in step 1 to the culture material in step 2, and then fully stir for 10-15 minutes to fully mix the bacteria in the culture material, adjust the culture temperature at 26-30 degrees Celsius, and culture for 5- For 10 days, add water to soak and rinse, and obtain a suspension after multiple washings;
- Step 4 Dilute the suspension in Step 3 to obtain a bacterial strain dilution
- Step 5 Take 10 parts by weight of sawdust, 10 parts of corn stalk chips, and 10 parts of wheat chips and add them to the mixer for stirring, so as to make them fully and evenly mixed, then add quicklime, potassium dihydrogen phosphate, magnesium sulfate and urea, and carry out Stir and mix again to obtain the propagation material;
- Step 6 Add the strain dilution in step 4 into the propagation material in step 5 at 1:10, then add sterile water, and mix to control the humidity between 55-65% and the temperature at 25- 30 degrees Celsius, the fermentation is stopped for 8-15 days, and it can be stored after air-drying.
- the suspension in the step 3 can be obtained by performing auxiliary treatment in a centrifuge.
- the washing solution is added to the centrifuge for medium-low speed centrifugation, and the upper layer liquid can be taken.
- the suspension in step 4 is diluted by adding sterile water, and the diluted concentration is 1 ⁇ 10 7 cfu/ml.
- the mass fractions of quicklime, potassium dihydrogen phosphate, magnesium sulfate and urea added in the step 5 are 1 part, 0.5 part, 0.2 part and 0.8 part respectively.
- the mass fractions of quicklime, potassium dihydrogen phosphate, magnesium sulfate and urea added in the step 5 are 1.2 parts, 0.3 parts, 0.1 parts and 0.9 parts respectively.
- the mass fractions of quicklime, potassium dihydrogen phosphate, magnesium sulfate and urea added in the step 5 are 0.8 part, 0.7 part, 0.3 part and 0.7 part respectively.
- the beneficial effect of the present invention is that a method for producing bacterial fertilizer by the propagation of Trichoderma of the present invention adopts cheap production raw materials, the operation is simple, no large-scale equipment is required, the industry is simple, and the produced Trichoderma yield is large,
- the method has high efficiency and is easy to popularize, and the method can be implemented on a large scale in stages, the obtained intermediate product suspension mixture can be stored as an intermediate product, and it is very convenient to use, and the produced Trichoderma can assist the growth of plants and improve the product of agricultural products.
- the invention discloses a method for the propagation of Trichoderma to produce bacterial fertilizer, comprising the following steps:
- Step 1 inoculate the Trichoderma strain into the PDA medium and cultivate for 3-6 days to obtain the Trichoderma species;
- Step 2 Take the weight fractions of 10 parts of corn flour, 10 parts of wheat bran, 5 parts of bran, 2 parts of sucrose, 1 part of calcium carbonate and 0.1 part of ferrous phosphate, add sterile water, and add them together in a mixer to fully stir and mix to prepare the mixture. into a liquid fermentation broth, adjust the pH to 5.5-6, and then squeeze out excess water, under high pressure and high temperature environment, carry out sterilization treatment for 30-40min, and naturally cool to obtain culture material;
- Step 3 Add the Trichoderma species in step 1 to the culture material in step 2, and then fully stir for 10-15 minutes to fully mix the bacteria in the culture material, adjust the culture temperature at 26-30 degrees Celsius, and culture for 5- For 10 days, add water to soak and rinse, and obtain a suspension after multiple washings;
- Step 4 Dilute the suspension in Step 3 to obtain a bacterial strain dilution
- Step 5 Take 10 parts by weight of sawdust, 10 parts of corn stalk chips, and 10 parts of wheat chips and add them to the mixer for stirring, so as to make them fully and evenly mixed, then add quicklime, potassium dihydrogen phosphate, magnesium sulfate and urea, and carry out Stir and mix again to obtain the propagation material;
- Step 6 Add the strain dilution in step 4 into the propagation material in step 5 at 1:10, then add sterile water, and mix to control the humidity between 55-65% and the temperature at 25- 30 degrees Celsius, the fermentation is stopped for 8-15 days, and it can be stored after air-drying.
- the suspension in the step 3 can be obtained by performing auxiliary treatment in a centrifuge.
- the washing solution is added to the centrifuge for medium-low speed centrifugation, and the upper layer liquid can be taken.
- the suspension is diluted by adding sterile water, and the diluted concentration is 1 ⁇ 10 7 cfu/ml.
- the mass fractions of quicklime, potassium dihydrogen phosphate, magnesium sulfate and urea added in the described step 5 are respectively 1 part, 0.5 part, 0.2 part and 0.8 part.
- the method for producing bacterial fertilizer by propagation of Trichoderma adopts cheap production raw materials, simple operation, no need for large-scale equipment, simple industry, large yield of Trichoderma produced, high efficiency, easy popularization, and the method can realize large-scale staged Implementation, the obtained intermediate product suspension mixture can be stored as an intermediate product, and is very convenient to use, and the produced Trichoderma can assist the growth of plants and improve the product of agricultural products.
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Abstract
一种木霉菌扩繁生产菌肥的方法，将木霉菌株接种到PDA培养基中培养3-6天，得到木霉菌种，按重量分数取玉米粉10份、麦糠10份、麸皮5份、蔗糖2份、碳酸钙1份以及磷酸亚铁0.1份加入无菌水，共同加入搅拌机中进行充分搅拌混合，制成液体发酵液，通过一系列工艺方法得到菌肥，该种木霉菌扩繁生产菌肥的方法，采用廉价的生产原料，操作简单，无需大型设备，工艺简单，生产的木霉菌产量大，效率高，容易推广，且该方法可以实现大规模分阶段实施，得到的中间产物悬浮混合液，可作为中间产物保存，使用极其方便，生产的木霉菌可以辅助植物生长。
Description
本发明涉及生物菌生产技术领域，尤其涉及一种木霉菌扩繁生产菌肥的方法。
木霉菌属真菌门，半知菌亚门，丝孢纲，丝孢目，丛梗孢科，木霉属，广泛存在于不同环境条件下的土壤中。自19世纪中叶，人类对木霉菌已有了初步的认识，但直到上世纪60年代木霉菌的分类地位才得以确定。大多数木霉菌可产生多种对植物病原真菌、细菌及昆虫具有拮抗作用的生物活性物质，比如细胞壁降解酶类和次级代谢产物，并能提高农作物的抗逆性，促进植物生长和提高农产品产量，因此被广泛用于生物防治、生物肥料及土壤改良剂。由于化学农药对环境的负面影响较为严重，所以对环境较为友好的生物农药木霉菌受到了广泛的关注。
为快速高效的得到大量的木霉菌，同时降低生产的经济成本，为此我们提供了一种木霉菌扩繁生产菌肥的方法。
发明内容
本发明的目的在于提供一种木霉菌扩繁生产菌肥的方法，以解决上述背景技术中提出的问题。
本发明的目的是通过下述技术方案予以实现：一种木霉菌扩繁生产菌肥的方法，包括以下步骤：
步骤1：将木霉菌株接种到PDA培养基中培养3-6天，得到木霉菌种；
步骤2：取重量分数玉米粉10份、麦糠10份、麸皮5份、蔗糖2份、碳酸钙1份以及磷酸亚铁0.1份加入无菌水，共同加入搅拌机中进行充分搅拌混合，制成液体发酵液，调整PH为5.5-6，然后挤出多余的水分，在高压高温环境下，进行灭菌处理30-40min，自然冷却得到培养料；
步骤3：将步骤1中的木霉菌种加入步骤2中的培养料中，然后充分搅拌10-15分钟，使菌种在培养料中充分混合，调节培养温度在26-30摄氏度，培养5-10天，加入清水浸泡冲洗，经过多次洗涤后得到悬浮液；
步骤4：将步骤3中的悬浮液进行稀释，得到菌种稀释液，
步骤5：取1重量份数木屑10份、玉米杆屑10份、麦屑10份共同加入搅拌机中进行搅拌，使其充分均匀混合，在加入生石灰、磷酸二氢钾、硫酸镁以及尿素，进行再次搅拌混合，得到扩繁料；
步骤6：将步骤4中的菌种稀释液按照1:10加入步骤5中的扩繁料中，然后加入无菌水，进行混合使其湿度控制在55-65％之间，温度为25-30摄氏度，发酵8-15天停止，风干后储存即可。
本发明中，所述步骤3中的悬浮液或通过离心机进行辅助处理得到，将洗涤液加入离心机中进行中低速离心处理，取上层液即可。
本发明中，所述步骤4中悬浮液通过加入无菌水进行稀释，稀释后的浓度为1X107cfu/ml。
本发明中，所述步骤5中加入的生石灰、磷酸二氢钾、硫酸镁以及尿素的质量份数分别为1份、0.5份、0.2份以及0.8份。
作为优选方案,所述步骤5中加入的生石灰、磷酸二氢钾、硫酸镁以及尿素的质量份数分别为1.2份、0.3份、0.1份以及0.9份。
作为优选方案,所述步骤5中加入的生石灰、磷酸二氢钾、硫酸镁以及尿素的质量份数分别为0.8份、0.7份、0.3份以及0.7份。
与现有技术相比，本发明的有益效果是该发明一种木霉菌扩繁生产菌肥的方法，采用廉价的生产原料，操作简单，无需大型设备，工业简单，生产的木霉菌产量大，效率高，容易推广，且该方法可以实现大规模分阶段实施，得到的中间产物悬浮混合液，可作为中间产物保存，使用及其方便，生产的木霉菌可以辅助植物生长，提高农产品的产品。
下面将结合本发明实施例中的附图，对本发明实施例中的技术方案进行清楚、完整地描述，显然，所描述的实施例仅仅是本发明一部分实施例，而不是全部的实施例，基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。
本发明公开了一种木霉菌扩繁生产菌肥的方法，包括以下步骤：
步骤1：将木霉菌株接种到PDA培养基中培养3-6天，得到木霉菌种；
步骤2：取重量分数玉米粉10份、麦糠10份、麸皮5份、蔗糖2份、碳酸钙1份以及磷酸亚铁0.1份加入无菌水，共同加入搅拌机中进行充分搅拌混合，制成液体发酵液，调整PH为5.5-6，然后挤出多余的水分，在高压高温环境下，进行灭菌处理30-40min，自然冷却得到培养料；
步骤3：将步骤1中的木霉菌种加入步骤2中的培养料中，然后充分搅拌10-15分钟，使菌种在培养料中充分混合，调节培养温度在26-30摄氏度，培养5-10天，加入清水浸泡冲洗，经过多次洗涤后得到悬浮液；
步骤4：将步骤3中的悬浮液进行稀释，得到菌种稀释液，
步骤5：取1重量份数木屑10份、玉米杆屑10份、麦屑10份共同加入搅拌机中进行搅拌，使其充分均匀混合，在加入生石灰、磷酸二氢钾、硫酸镁以及尿素，进行再次搅拌混合，得到扩繁料；
步骤6：将步骤4中的菌种稀释液按照1:10加入步骤5中的扩繁料中，然后加入无菌水，进行混合使其湿度控制在55-65％之间，温度为25-30摄氏度，发酵8-15天停止，风干后储存即可。
本发明中，所述步骤3中的悬浮液或通过离心机进行辅助处理得到，将洗涤液加入离心机中进行中低速离心处理，取上层液即可。所述步骤4中悬浮液通过加入无菌水进行稀释，稀释后的浓度为1X107cfu/ml。所述步骤5中 加入的生石灰、磷酸二氢钾、硫酸镁以及尿素的质量份数分别为1份、0.5份、0.2份以及0.8份。
该种木霉菌扩繁生产菌肥的方法，采用廉价的生产原料，操作简单，无需大型设备，工业简单，生产的木霉菌产量大，效率高，容易推广，且该方法可以实现大规模分阶段实施，得到的中间产物悬浮混合液，可作为中间产物保存，使用及其方便，生产的木霉菌可以辅助植物生长，提高农产品的产品。
对于本领域技术人员而言，显然本发明不限于上述示范性实施例的细节，而且在不背离本发明的精神或基本特征的情况下，能够以其他的具体形式实现本发明。因此，无论从哪一点来看，均应将实施例看作是示范性的，而且是非限制性的，本发明的范围由所附权利要求而不是上述说明限定，因此旨在将落在权利要求的等同要件的含义和范围内的所有变化囊括在本发明内。不应将权利要求中的任何附图标记视为限制所涉及的权利要求。
此外，应当理解，虽然本说明书按照实施方式加以描述，但并非每个实施方式仅包含一个独立的技术方案，说明书的这种叙述方式仅仅是为清楚起见，本领域技术人员应当将说明书作为一个整体，各实施例中的技术方案也可以经适当组合，形成本领域技术人员可以理解的其他实施方式。
Claims (6)
- 一种木霉菌扩繁生产菌肥的方法，其特征在于：包括以下步骤：步骤1：将木霉菌株接种到PDA培养基中培养3-6天，得到木霉菌种；步骤2：取重量分数玉米粉10份、麦糠10份、麸皮5份、蔗糖2份、碳酸钙1份以及磷酸亚铁0.1份加入无菌水，共同加入搅拌机中进行充分搅拌混合，制成液体发酵液，调整PH为5.5-6，然后挤出多余的水分，在高压高温环境下，进行灭菌处理30-40min，自然冷却得到培养料；步骤3：将步骤1中的木霉菌种加入步骤2中的培养料中，然后充分搅拌10-15分钟，使菌种在培养料中充分混合，调节培养温度在26-30摄氏度，培养5-10天，加入清水浸泡冲洗，经过多次洗涤后得到悬浮液；步骤4：将步骤3中的悬浮液进行稀释，得到菌种稀释液，步骤5：取1重量份数木屑10份、玉米杆屑10份、麦屑10份共同加入搅拌机中进行搅拌，使其充分均匀混合，在加入生石灰、磷酸二氢钾、硫酸镁以及尿素，进行再次搅拌混合，得到扩繁料；步骤6：将步骤4中的菌种稀释液按照1:10加入步骤5中的扩繁料中，然后加入无菌水，进行混合使其湿度控制在55-65％之间，温度为25-30摄氏度，发酵8-15天停止，风干后储存即可。
- 根据权利要求1所述的一种木霉菌扩繁生产菌肥的方法，其特征在于：所述步骤3中的悬浮液或通过离心机进行辅助处理得到，将洗涤液加入离心机中进行中低速离心处理，取上层液即可。
- 根据权利要求1所述的一种木霉菌扩繁生产菌肥的方法，其特征在于：所述步骤4中悬浮液通过加入无菌水进行稀释，稀释后的浓度为1X107cfu/ml。
- 根据权利要求1所述的一种木霉菌扩繁生产菌肥的方法，其特征在于：所述步骤5中加入的生石灰、磷酸二氢钾、硫酸镁以及尿素的质量份数分别为1份、0.5份、0.2份以及0.8份。
- 根据权利要求1所述的一种木霉菌扩繁生产菌肥的方法，其特征在于： 所述步骤5中加入的生石灰、磷酸二氢钾、硫酸镁以及尿素的质量份数分别为1.2份、0.3份、0.1份以及0.9份。
- 根据权利要求1所述的一种木霉菌扩繁生产菌肥的方法，其特征在于：所述步骤5中加入的生石灰、磷酸二氢钾、硫酸镁以及尿素的质量份数分别为0.8份、0.7份、0.3份以及0.7份。
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