Simple and convenient separation method for phytophthora parasitica var nicotianae

Abstract

The invention provides a simple and convenient separation method for phytophthora parasitica var nicotianae. By the method, a clean working environment is built by using simple equipment and spatial conditions so as to fast separate the phytophthora parasitica var nicotianae. The method particularly comprises the following steps of: (1) selecting and collecting diseased or infected plants; (2) purifying working environment; (3) separating germs; and (4) culturing the germs at room temperature; and the like. Namely, the method comprises the following steps of: building the clean working environment under simple conditions; sterilizing the working environment by the flame of an alcohol burner to form a sterile microenvironment; dissecting diseased or infected stems; picking up disk-shaped diseased or infected medullary slices in the diseased or infected stems with tweezers; placing the medullary slices on oat or lima bean medium plates, wherein the medium plates are sealed by sealing films, and 3 to 5 plates are arranged in a sealed culture box; and culturing the medullary slices for 3 to 5 days at room temperature to obtain target germs by colonial morphology. The method is suitable for field sampling and timely separating, uses simple equipment in the whole operation, and can separate the phytophthora parasitica var nicotianae without microbiology laboratories. The separated germs have high purity and the characteristics of germ wild types.

Description

technical field

[0001] The invention belongs to the technical field of separation of agricultural microorganisms, and in particular relates to a simple separation method for efficiently and rapidly separating tobacco black shank bacteria using simple equipment or conditions. Background technique

[0002] Tobacco black shank (tobacco black shank) is one of the most serious diseases in tobacco production in the world, especially in temperate, subtropical and tropical regions. It is also one of the main diseases of tobacco in my country. The isolation method of tobacco black shank pathogen is usually to collect diseased strains in diseased fields, and utilizes selective culture medium plate to isolate pathogenic bacteria (Zheng Xiaobo, Phytophthora and its research technology, Beijing, China Agricultural Press, 1997, P84 in laboratory) -85). Therefore, there is often a time difference of 3 to 7 days or longer between the collection of the disease sample and the isolation of th...

Examples

Embodiment 1

[0025] Select the diseased plants with typical symptoms in the field, no other symptoms, no cracking or artificial dissection of the diseased stems, that is, the diseased plants with the diseased pith discs sealed and wrapped in healthy stems as the target diseased plants, and remove the stem tops when collecting , leaves and roots, only keep the diseased stems, wash the surface to remove floating dust, and dry them. Each diseased stem is wrapped and separated with sterilized toilet paper, and placed in a disposable clean sealed bag. To isolate the pathogen in a clean environment, first cut the diseased stem from the upper part of the stem, grasp the upper end of the diseased stem with both hands, and pull the diseased stem until the disc-shaped diseased pith is exposed. Then use flame-sterilized elbow tweezers to pick up the lesion slices and place them in the oat culture medium plate under the flame; The time for placing the slices was 3 to 5 seconds; each sampling point was...

Embodiment 2

[0027] Replace the oat medium plate with the lima bean medium plate, and the others are the same as in Example 1.

Embodiment 3

[0029] Select diseased stems in the manner of Example 1, separate diseased marrow slices and carry out room temperature cultivation, and select target pathogenic bacteria according to the pure white colony form. The pathogenicity of the isolated bacteria was verified by inoculating the susceptible variety Honghua Dajinyuan with the Jungu method, and the strains with a disease severity greater than 1 were pathogenic.

technical field

[0001] The invention belongs to the technical field of separation of agricultural microorganisms, and in particular relates to a simple separation method for efficiently and rapidly separating tobacco black shank bacteria using simple equipment or conditions. Background technique

[0002] Tobacco black shank (tobacco black shank) is one of the most serious diseases in tobacco production in the world, especially in temperate, subtropical and tropical regions. It is also one of the main diseases of tobacco in my country. The isolation method of tobacco black shank pathogen is usually to collect diseased strains in diseased fields, and utilizes selective culture medium plate to isolate pathogenic bacteria (Zheng Xiaobo, Phytophthora and its research technology, Beijing, China Agricultural Press, 1997, P84 in laboratory) -85). Therefore, there is often a time difference of 3 to 7 days or longer between the collection of the disease sample and the isolation of th...

Examples

Embodiment 1

[0025] Select the diseased plants with typical symptoms in the field, no other symptoms, no cracking or artificial dissection of the diseased stems, that is, the diseased plants with the diseased pith discs sealed and wrapped in healthy stems as the target diseased plants, and remove the stem tops when collecting , leaves and roots, only keep the diseased stems, wash the surface to remove floating dust, and dry them. Each diseased stem is wrapped and separated with sterilized toilet paper, and placed in a disposable clean sealed bag. To isolate the pathogen in a clean environment, first cut the diseased stem from the upper part of the stem, grasp the upper end of the diseased stem with both hands, and pull the diseased stem until the disc-shaped diseased pith is exposed. Then use flame-sterilized elbow tweezers to pick up the lesion slices and place them in the oat culture medium plate under the flame; The time for placing the slices was 3 to 5 seconds; each sampling point was...

Embodiment 2

[0027] Replace the oat medium plate with the lima bean medium plate, and the others are the same as in Example 1.

Embodiment 3

[0029] Select diseased stems in the manner of Example 1, separate diseased marrow slices and carry out room temperature cultivation, and select target pathogenic bacteria according to the pure white colony form. The pathogenicity of the isolated bacteria was verified by inoculating the susceptible variety Honghua Dajinyuan with the Jungu method, and the strains with a disease severity greater than 1 were pathogenic.