Materials and utensils: specimen holder, absorbent paper, plastic bag, paper bag, label, pencil, marker, knife, scissors, hand saw, specimen jar, copper glycolate, copper sulfate, 95% ethanol, formaldehyde solution, sulfurous acid, glycerol, distilled water, etc.
Contents and methods
(1) Collection of disease specimens
1 Basic requirements for specimen collection
To collect a complete specimen, it is best to take photos first, and then press or soak them for preservation. The clearer the picture, the better. Normal plants and diseased parts need to be photographed. For each specimen, at least 2-3 samples should be collected.
2 Sampling location
(1) If there are fruiting bodies on the specimen, try to collect them on the old leaves.
Because it is mature, the fruiting body of many fungi in sexual stage appears on litter, while the fruiting body in asexual stage can be found in living body.
(2) For soft and juicy fruiting bodies or fruit materials, newly diseased young fruits should be collected.
(3) The virus disease should try to collect the top and new leaves.
(4) The diseased tissues should be collected for nematodiasis specimens, and the soil around the root system should be collected for nematodiasis specimens that harm the root system.
(5) Wilting plants should be dug up by roots, and sometimes they should be collected with rhizosphere soil.
(6) For thicker branches and plants, it is advisable to cut a piece or cut a piece.
(7) If the diseases appear on leaves, fruits and branches, try to collect them comprehensively.
(8) Parasites should be collected together with the host. Diseases with special symptoms should be collected with branches, leaves or flowers of plants in order to identify and determine their host names.
3 specimen data record and specimen number
Records shall be accurate, concise and complete. A complete record and label are equally important, host name,
Date and place of collection, name of collector, ecological conditions and soil conditions, etc.
4 specimen collection method
(1) Collect disease specimens of branches and roots.
Use a saw, branch scissors or high branch scissors to remove the affected part. Don't break it by hand, it will affect the beauty of the specimen. Pay special attention to the long and strong branches of fibers.
(2) Collection of leaf disease specimens
Large-leaved plants, whose leaves and inflorescences are very large, can be collected in part or in sections when collecting specimens. A specimen consists of young leaves and flowers on the same plant (with the actual size of leaves indicated); Or take a part of leaves and petioles respectively;
Parasitic plants such as Cuscuta chinensis and Leguminosae should be collected together with the host, and the host name should be recorded. (3) Collection of disease specimens of succulent or juicy plants (flowers and fruits)
To cut vertically or horizontally, sometimes it is necessary to dig out the internal tissue, and it is also necessary to consider whether to soak half of the materials in a preservation solution for preservation. When drying in the field, a large amount of salt should be sprinkled on the surface of the cut stem, and the materials wrapped with salt should be placed in a specimen folder with multiple newspapers. After 24 hours, the newspaper soaked in salt water should be taken out, or the material should be scalded to death with boiling water, and then put into the cardboard-aluminum interlayer to dry.
(2) Preparation of disease specimens
Except for some samples collected from the field for isolation and identification, it is best to take pictures of typical disease symptoms first, and then press or soak them for preservation. The pressed or impregnated specimens keep their original properties as much as possible, and tiny specimens can be made into slides, such as double-layer slides, concave slides or collected in other small glass tubes.
(3) Preservation of disease specimens
1 drying method is suitable for making specimens of stem, leaf and other diseases, which requires less water, is the simplest, most economical and most widely used. The collected specimens are put in absorbent paper, and at the same time, labels with the collection place, date and host name are put in. Then press it with a specimen holder and bake or heat it in the sun to dry it. The faster they dry, the better their original color and the higher their quality. Specimens collected in summer are prone to mildew and discoloration under high temperature and high humidity, so it is advisable to change paper frequently. Usually, the paper is changed L-2 times a day in the first three or four days (depending on the water content and temperature and humidity of the specimen), and then every two or three days until it is completely dry. When changing the paper for the first time, you should sort out the specimens, because the specimens become soft and easy to spread after preliminary drying. The stems and leaves of tobacco, broad beans, pears and potatoes are easy to turn black and discolor, and the specimens are not easy to preserve color. Special attention should be paid to rapid drying in the production process. If it is necessary to keep the specimen green and dry, the specimen can be soaked in 2-4% copper sulfate solution for 24 hours, or treated with copper acetate solution before pressing (see impregnation method of copper acetate in impregnation method for formula and method). You can also apply a layer of liquid paraffin to the leaves, and then press them to keep them bright green for 2-3 days.
2 soaking method is suitable for meat disease specimens,
The fruits, tubers or fruiting bodies of basidiomycetes must be preserved by soaking. There are many kinds of immersion liquids, some of which are purely antiseptic, and some are used to keep the original color of the specimen. This paper introduces several common and effective methods.
(1) Anti-corrosion impregnation method: This impregnation method can only prevent corrosion but not preserve color, such as radish and sweet potato. After cleaning, color-preserving specimens are not required to be directly immersed in the following solution. ① 5% formalin impregnation solution.
② Sulfuric acid impregnation solution: add 5%-6% sulfurous acid15ml in1000ml of water (generally, the sulfurous acid content in the market is 5-6%) or use10.5g of sodium sulfite, 8ml of concentrated sulfuric acid and 500ml of water to prepare a mixed solution.
(2) Method of soaking solution and preserving green specimens:
(1) copper acetate color-preserving impregnation method: gradually add copper acetate crystals into 50% acetic acid and dissolve them to saturation as mother liquor (adding 15g copper acetate can make 50% acetic acid about 1000ml saturated), and dilute with water 1-4 times when using (the dilution multiple depends on the color of the specimen, and the color. Boil the solution and put it into the specimen. First, the green color of the specimen was removed, and then the specimen returned to green after a few minutes. At this time, take out the specimen immediately, rinse it with clear water and store it in 5% formalin solution. This method is called heat treatment. The method of cold treatment is to soak the specimen in the above diluent for 3 hours, then the specimen will fade and return to green after 72 hours. At this point, the specimen will be taken out, washed with water and stored in 5% formalin solution. This method has a long color retention time, and its color retention principle is roughly that copper ions in chlorophyll replace magnesium ions. Therefore, after the solution has been treated for many times, copper ions will gradually decrease. If you want to continue to use it, you should add an appropriate amount of copper acetate. Specimens preserved in this way are often slightly blue, which is slightly different from the original color of plant specimens.
(2) Color-preserving impregnation method of copper sulfate: rinse the specimen with clear water and directly soak it in 5% copper sulfate solution 1-7 days. When the specimen is brown, take it out and rinse it with clear water to remove the excess copper sulfate solution on the surface of the specimen, and then store it in sulfurous acid impregnation solution.
(3) Preserve the soaking solution of yellow and orange samples: preserve the samples of apricot, pear, persimmon, red pepper and other fruits, and soak them with sulfurous acid.
Pickling solution and sulfurous acid impregnation solution have bleaching effect, so pay attention to the concentration when using, generally 1% (the concentration can be thinner when soaking apricots). Because the concentration is too small and the corrosion resistance is not enough, a proper amount of alcohol can be added. In order to prevent the fruit from cracking, you can add a little glycerin.
(4) Impregnation solution for preserving red specimens: The red color of specimens is difficult to preserve, because red is a water-soluble anthocyanin, which is difficult to preserve. It is usually preserved in hessler immersion liquid, and its components are: 50g of zinc chloride, 25ml of formalin, 25ml of glycerol, and water 1000ml. Dissolve zinc chloride in hot water, add formalin, and use its clear solution if there is precipitation. The solution is suitable for samples that are red due to anthocyanins, such as apples and tomatoes.
(4) Preservation of specimens
The prepared specimens are classified and registered, and then sorted and preserved according to a certain system.
Fungal specimens are generally arranged according to the classification system, and there should be two sets of index cards, one is the host index and the other is the fungus index, which is convenient for searching, dragging and sorting.
The specimen room (cabinet) should be kept dry and spacious, and applied regularly to prevent moth and mildew.
1 specimen box is more convenient for storing dry disease specimens for teaching and demonstration, and the suitable size of glass carton is 20×28×2 cm. First spread a layer of cotton in the carton, and add a little camphor powder to the cotton to repel insects. Finally, put samples and labels on cotton.
2 specimen bottle preservation The impregnated specimen is kept in the specimen bottle. In order to prevent the specimen from sinking and floating, the specimen can be tied to a glass strip and then put into a specimen bottle. After the specimen bottle cap is closed, add paraffin to seal it, and then label it.
The paper is folded with kraft paper to form a paper sleeve of 15×33cm, and most of the dried specimens are kept in the paper sleeve. Put the specimen in a paper sleeve, put a label on the paper sleeve and put it in the specimen cabinet.