Nitrogen-fixing microorganism G96 as well as bacterium agent preparation method and application thereof

A nitrogen-fixing microorganism and inoculum technology, applied in the field of nitrogen-fixing microorganism G96 and the preparation of inoculants

Active Publication Date: 2016-10-12
DONGGUAN BAODE BIOLOGICAL ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because Bacillus has many advantages such as storage resistance, strong stress resistance and salt-alkali tolerance, but at the same time, relatively few nitrogen-fixing bacteria can produce spores found in current research, so the selection of Bacillus strains with nitrogen-fixing function is very important for microbial fertilizers. production is important

Method used

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  • Nitrogen-fixing microorganism G96 as well as bacterium agent preparation method and application thereof
  • Nitrogen-fixing microorganism G96 as well as bacterium agent preparation method and application thereof
  • Nitrogen-fixing microorganism G96 as well as bacterium agent preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] A nitrogen-fixing microorganism G96 in this embodiment, the nitrogen-fixing microorganism G96 is preserved in the China Center for Type Culture Collection, and the preservation number is CCTCCM 2015752. The nitrogen-fixing microorganism G96 has the DNA sequence of No. 1 in the sequence table.

[0083] The nitrogenase activity of the nitrogen-fixing microorganism G96 was 1050.56 nmol / (mL·h), and the secretion of indole acetic acid during the growth and metabolism process was 356.18 mg / L.

[0084] A method for preparing a nitrogen-fixing microorganism G96 bacterial agent, comprising the following steps:

[0085] (1) Separation and screening

[0086] Select soil samples containing nitrogen-fixing bacteria colonies of nitrogen-fixing microorganism G96, and obtain nitrogen-fixing bacteria colonies through isolation and screening medium culture;

[0087] (2) Purification and preservation

[0088] Purify the isolated and screened nitrogen-fixing bacteria colony on the purif...

Embodiment 2

[0099] The difference between this embodiment and embodiment 1 is that the following steps are also included between the step (2) and step (3) of this embodiment:

[0100] (S1) Gram staining: perform Gram staining on the purified single colony, and screen to obtain positive bacteria;

[0101] (S2) Spore staining: the positive bacteria are stained with spores, and a single colony of Gram-positive bacteria containing spores is obtained by screening.

[0102] Specifically, the Gram staining method is:

[0103] (1) Smear: In a sterile operating table, take a glass slide and bake it slightly above the flame lamp to remove impurities on the slide. Drop a drop of sterile water in the center of the slide, pick a single colony in the water drop, and spread evenly with a burnt inoculation loop. Pass the sample slide back and forth 3 times over the fire lamp to fix the cells.

[0104] (2) Initial dyeing: Add 2-5 drops of ammonium oxalate crystal violet dye solution, dye for 1 min, pou...

Embodiment 3

[0113] The difference between this example and Example 1 or 2 is that the nitrogenase activity of the nitrogen-fixing microorganism G96 in this example is 1000nmol / (mL h), and the secretion amount of indole acetic acid during its growth and metabolism is 300mg / L.

[0114] In the step (1), the separation and screening medium is made from raw materials of the following quality: CaCO 3 1.0g, MgSO 4 ·7H 2 O 0.6g, K 2 HPO 4 1.0g, NaCl 0.1g, FeSO 4 ·7H 2 O 0.001g, NaMO 4 2H 2 O 0.05g, sucrose 5g, agar 18g, distilled water 1000ml, the pH of described separation and screening medium is 7.2.

[0115] The specific method of purification and preservation in the step (2) is as follows: Streak and purify the isolated and screened colonies on the purification and preservation medium, culture at a constant temperature at 30°C for 38 hours to separate and obtain a single colony of nitrogen-fixing microorganism G96, and remove the colonies that appear on the plate. The single coloni...

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Abstract

The invention relates to the technical field of nitrogen-fixing microorganism bacterium agents and in particular relates to a nitrogen-fixing microorganism G96 as well as a bacterium agent preparation method and application thereof. The nitrogen-fixing microorganism G96 is preserved in China Center for Type Culture Collection, with the preservation number of CCTCCM 2015752. The bacterium agent preparation method of the nitrogen-fixing microorganism comprises the following steps: (1) separating and screening; (2) purifying and preserving; (3) carrying out solid fermentation culture; (4) preparing the bacterium agent to obtain a bacillus megaterium G96 bacterium agent. The nitrogen-fixing microorganism G96 not only has relatively high nitrogenase activity, but also has a very high IAA (Indole Acetic Acid) secretion capability; the nitrogenase activity is 1000nmol / (mL h) to 1100nmol / (Ml h); the secretion amount of the IAA is 300mg / L to 400mg / L in a growth and anabolism process of the nitrogen-fixing microorganism.

Description

technical field [0001] The invention relates to the technical field of nitrogen-fixing microbial bacterial agents, in particular to a nitrogen-fixing microorganism G96 and a preparation method and application of the bacterial agent. Background technique [0002] Microbial fertilizer is a new type of fertilizer developed by utilizing the phosphorus-dissolving, potassium-dissolving and nitrogen-fixing properties of microorganisms. It can not only increase crop production and improve the quality of agricultural products, but also improve soil and increase soil fertility. Nitrogen-fixing microbial fertilizer is a new type of fertilizer that is energy-saving and environmentally friendly and meets the requirements of sustainable ecological agriculture. It fixes N in the air through nitrogen-fixing microorganisms. 2 , and convert it into NH for direct use by crops 4 + , so as to achieve the purpose of reducing the amount of chemical fertilizers. Because Bacillus has many advanta...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A01N63/02A01P21/00C12R1/11
CPCA01N63/10C12N1/205C12R2001/11
Inventor 陈三凤杨国平林敏孙旭生王亚君杨盼盼尹坤张学贤沈世华谭志远燕永亮
Owner DONGGUAN BAODE BIOLOGICAL ENG
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