A dse fungus and a method for rapid mycorrhization of blueberry tissue cultured seedlings

A technology of tissue culture seedlings and fungi, applied in the field of microorganisms, can solve the problems of low infection efficiency, long co-cultivation period, slow growth of DSE bacteria sheets, etc.

Active Publication Date: 2020-05-26
LUDONG UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, during the implementation of this method, DSE flakes often grow slowly in the plant medium, and the plant roots cannot fully contact with DSE, resulting in a long co-cultivation period and low infection efficiency.
Therefore, the above-mentioned method is not suitable for the symbiotic cultivation of slender and underdeveloped plants such as blueberry and DSE

Method used

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  • A dse fungus and a method for rapid mycorrhization of blueberry tissue cultured seedlings
  • A dse fungus and a method for rapid mycorrhization of blueberry tissue cultured seedlings
  • A dse fungus and a method for rapid mycorrhization of blueberry tissue cultured seedlings

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Experimental program
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Effect test

Embodiment 1

[0030] The isolation and purification of blueberry root endophytic fungi are as follows:

[0031] 1. Wash the blueberry root section with clean water, and then transfer it to an ultra-clean workbench for the following operations: soak the root section in 10% (mass ratio) hydrogen peroxide for 8 minutes, and gently turn it several times with sterile tweezers to Ensure thorough disinfection; after rinsing with sterile water for 2-3 times, turn it into 6% (mass ratio) sodium hypochlorite, soak for 15 minutes, and rinse with sterile water for 2 times; filter paper absorbs the residual liquid on the surface of the root segment. The sterilized root segments were cut into 0.5 cm long root segments with sterile scissors, inserted into the prepared PDA plate, and cultured upside down in the dark in a constant temperature incubator at 25°C. Recipe of PDA plate: 0.6% potato dipping powder, 2% glucose, 2% agar, pH5.6. Sterilize at 115°C for 20min, when cooled to 60°C, add sterile ampicill...

Embodiment 2

[0033] Morphological and molecular biology identification of embodiment 2 bacterial strain R16

[0034] 1. Pick the above-mentioned preserved purified strains, inoculate them on PDA plates, and culture and activate them in a constant temperature incubator at 25°C for 1-2 weeks. Use a sterile puncher with a diameter of 0.5 cm to take the bacteria cake from the outermost layer of the colony, inoculate it on a new PDA plate, and culture it in a constant temperature incubator at 25°C for 1 week to observe the shape of the colony. The R16 colony is off-white, tapetum-like, with smooth edges, and the back of the colony is gray-yellow ( figure 1 ).

[0035] 2. Use an inoculation needle to pick a small amount of purified single-colony hyphae, put it on a glass slide with a drop of normal saline to make a temporary mount, and observe the hyphae structure of the endophytic fungus under a microscope. The hyphae is bifurcated, and two side branches are issued from different planes by th...

Embodiment 3

[0039] The cultivation of embodiment 3 blueberry tissue culture seedlings, the steps are as follows:

[0040] (1) Differentiation of blueberry buds

[0041] Select blueberry tissue culture seedlings with good growth status and plant height of 4-5cm for rapid propagation. Use sterile scissors to cut off the blueberry plant in the tissue culture bottle from the base in an ultra-clean workbench, cut into 2-3cm stem segments, and then use sterile tweezers to connect it to the tissue culture bottle containing the differentiation medium , put 3-4 stem segments in each bottle, and spread them evenly. The medium used is based on WPM medium as the basic medium. In each liter of WPM medium, 20g of sucrose, 5-10g of agar, 1.0-3.0mg of phytohormone zeatin are added, and the pH of the medium is 5.0-5.5. The volume of the container is 10-30%, and it is autoclaved at 121°C for 20 minutes. The culture conditions are set at 23-25°C, light intensity of 2000-3000lx, light time of 8-16h / day, a...

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Abstract

A DSE fungus and a method for rapid mycorrhization of blueberry tissue cultured seedlings belong to the field of biotechnology. The strain is R16, and the preservation number is CGMCC No.13885. The invention also establishes a method for rapid mycorrhization of the bacterial strain and blueberry tissue cultured seedlings. The system uses sterilized moss as the co-cultivation substrate under sterile conditions, and the DSE inoculant adopts the liquid culture method. Compared with the current method of using bacterial block inoculation, the strain R16 inoculated with liquid culture grows faster, has sufficient contact with the blueberry rhizome, has a shorter co-cultivation time, and has a higher colonization rate. DSE strain R16 can significantly promote the growth of blueberry tissue culture seedlings. Compared with the non-inoculated control, the blueberry seedlings inoculated with DSE bacteria had significant advantages in growth, with large and dark green leaves, well-developed root system, and high transplanting survival rate.

Description

technical field [0001] The invention belongs to the field of microorganisms, and in particular relates to a DSE fungus and a method for rapid mycorrhization of blueberry tissue cultured seedlings. Background technique [0002] Dark septate endophytes (Dark Septate Endophytes, DSE) are a kind of mycorrhizal fungi that widely exist in plant roots and have dark septate hyphae. Existing studies have confirmed that DSE and the host have a mutualistic symbiosis, endow the plant with excellent growth traits, and can improve the host's ability to resist diseases and insect pests and stress resistance in stress environments. [0003] Blueberry (Vaccinium spp.), also known as lingonberry, is a plant of the genus Vaccinium in the Rhododendronaceae family. It is a new fruit tree species with high economic value and broad development prospects. The expansion of the cultivation area also increases the market demand for blueberry seedlings. The propagation of blueberry seedlings mainly ad...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/14A01N63/30A01P21/00A01G18/00A01G18/20A01G18/50C12R1/645
CPCA01G18/10A01N63/30C12N1/14C12N1/145C12R2001/645Y02A40/10
Inventor 宿红艳程显好宋方圆李媛媛孙志超曹思琪
Owner LUDONG UNIVERSITY
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