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Preparation method of improved fungus protoplasts

A protoplast and fungus technology, applied in the field of fungal protoplast preparation, can solve the problems of restricting the success rate, the number of protoplasts, the regeneration rate and the single nucleus rate, etc., and achieve high regeneration rate and single nucleus rate, and low pollution rate. Effect

Inactive Publication Date: 2019-12-27
GUANGDONG INST OF MICROBIOLOGY GUANGDONG DETECTION CENT OF MICROBIOLOGY +1
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Problems solved by technology

[0003] However, the author's research has found that only the "pollution rate" in the preparation method of fungal protoplasts in the prior art has seriously restricted the success rate of the preparation of protoplast monokaryotic strains. rate is relatively low

Method used

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  • Preparation method of improved fungus protoplasts
  • Preparation method of improved fungus protoplasts
  • Preparation method of improved fungus protoplasts

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Experimental program
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Embodiment 1

[0022] 1. Preparation of reagents and consumables required for the experiment

[0023] 1. The 0.22μm sterile filter membrane, centrifuge tube, disposable syringe (for filtration), pipette tip, disposable petri dish, and parafilm are all sterile.

[0024] 2. Medium with membrane: cut the cellophane membrane into a circle with a diameter of about 8 cm, and sterilize it for later use; pour the sterilized enriched comprehensive PDA medium into a flat plate (petri dish with a diameter of 9 cm) in the ultra-clean bench, and wait for After the medium plane is slightly solidified, a layer of sterilized cellophane film is spread on it to form a film-coated medium.

[0025] Enrichment of comprehensive PDA medium: first wash and peel the potatoes, then weigh 200g potatoes and cut them into small pieces, add water and boil them, filter to get the filtered juice, then add 20g of agar, 20g of glucose, KH 2 PO 4 3g, MgSO 4 Mix 1.5g, peptone 10g, and vitamin B1 0.3g, add water to a total v...

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Abstract

The invention discloses a preparation method of improved fungus protoplasts. Fungal mycelium is washed, a wall is broken by enzymatic hydrolysis, and protoplasts are collected for regeneration culture, finally, a target single colony is selected, and whether a single karyon is tested or not is conducted to obtain a protoplast mononuclear strain; and the preparation method of the fungal mycelium comprises the steps of covering a layer of sterilized membrane on a fungal solid culture medium to obtain a membrana-covered culture medium, inoculating the fungus on the membrana-covered culture medium, then culturing the fungus until the mycelium grows, taking the mycelium on the membrane, and then obtaining the fungal mycelium. According to the preparation method of the improved fungus protoplasts, the pollution rate is low, the number of the protoplasts, the regeneration rate and the mononuclear rate are high, and stable and efficient basic technical support for a fungal omics research and crossbreeding is provided.

Description

Technical field: [0001] The invention belongs to the field of fungal protoplast preparation, and in particular relates to an improved preparation method of fungal protoplast. Background technique: [0002] Protoplast preparation is a commonly used technical method for fungal researchers. It can be used to obtain monokaryons and provide high-quality test materials for myconomics research and hybrid breeding. Therefore, it is an extremely important basic technology. The most common method for the preparation of fungal protoplast monokaryotic strains reported in the literature at present is to first carry out liquid fermentation of the strain; then collect the liquid mycelia for repeated washing, enzymatic wall breaking (physical wall breaking was also used in the early days), and protoplast collection; Then carry out regeneration culture; finally pick the target single colony, and check whether it is monokaryon. The effect of protoplast preparation of monokaryon mainly depend...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12R1/645
CPCC12N1/14
Inventor 黄龙花史钏刘远超莫伟鹏梁晓薇胡惠萍
Owner GUANGDONG INST OF MICROBIOLOGY GUANGDONG DETECTION CENT OF MICROBIOLOGY