CN110320079B - Rhizopus slide specimen and manufacturing method thereof - Google Patents
Rhizopus slide specimen and manufacturing method thereof Download PDFInfo
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- CN110320079B CN110320079B CN201910637380.2A CN201910637380A CN110320079B CN 110320079 B CN110320079 B CN 110320079B CN 201910637380 A CN201910637380 A CN 201910637380A CN 110320079 B CN110320079 B CN 110320079B
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- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 18
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 12
- 239000003795 chemical substances by application Substances 0.000 claims description 11
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Images
Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/2813—Producing thin layers of samples on a substrate, e.g. smearing, spinning-on
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
-
- G—PHYSICS
- G09—EDUCATION; CRYPTOGRAPHY; DISPLAY; ADVERTISING; SEALS
- G09B—EDUCATIONAL OR DEMONSTRATION APPLIANCES; APPLIANCES FOR TEACHING, OR COMMUNICATING WITH, THE BLIND, DEAF OR MUTE; MODELS; PLANETARIA; GLOBES; MAPS; DIAGRAMS
- G09B23/00—Models for scientific, medical, or mathematical purposes, e.g. full-sized devices for demonstration purposes
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
- G01N2001/305—Fixative compositions
Abstract
The invention relates to a method for manufacturing a biological slide specimen, and discloses a rhizopus slide specimen and a method for manufacturing the rhizopus slide specimen. The invention also protects the rhizopus slide specimen prepared by the method for preparing the rhizopus slide specimen. The method for manufacturing the rhizopus slide specimen can clearly observe the sporangium and the rhizoid, and is simple and easy to implement and high in success rate.
Description
Technical Field
The invention relates to a method for manufacturing a biological slide specimen, in particular to a rhizopus slide specimen and a manufacturing method thereof.
Background
The rhizopus has wide distribution and is generally harmless to people, and the edible liqueur and the saccharified feed are prepared by selecting rhizopus strains with strong saccharifying power, but the wide distribution can cause laboratory pollution. It can also cause food mildew, is a conditional pathogen, can cause mucormycosis, is often accompanied by diabetes and acidosis, and easily invades nasal mucosa and submucosal tissue to cause cellulitis. It also can invade brain tissue to cause brain lesion. Invade blood vessels, cause embolism, etc. Therefore, rhizopus is also a zygomycete which is mainly explained in teaching and is a necessary object to be observed in mycology and microbiology experiments.
Traditional rhizopus is cultured on a Sabouraud's dextrose agar medium at room temperature, the growth is rapid, the hyphae are white, become grey or brownish black after a long time, are cotton-like colonies, are often filled in a slant test tube or a plate, and are thick in microscopic examination hyphae and are not separated or are rarely separated. Forming false root on the stolon hypha, growing a cyst handle in the direction opposite to the false root, and forming a sporangium at the top. The sporangia is spherical, and is initially white and then black, and spherical or oval cystospores are filled in the sporangia. Therefore, the important identification characteristics of rhizopus are sporangium and rhizoid, however, in the experimental teaching, the rhizopus slide purchased by people is usually cultured in a liquid state, in order to ensure sufficient oxygen supply, a stirring tank body or a shaking flask is needed for culture, so that thalli are damaged (sporangium and rhizoid fall off), and the target characteristics are difficult to observe by directly purchasing the rhizopus slide, so that the method for preparing the rhizopus slide specimen is provided.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide a rhizopus slide specimen and a manufacturing method thereof, which can clearly observe sporangium and rhizoid, and have the advantages of simple and easy method and high success rate.
The invention aims to provide a method for manufacturing a rhizopus slide specimen, which comprises the steps of placing a culture medium at the bottom of a first culture dish, inoculating rhizopus in the center of the bottom of the first culture dish, culturing at 25 +/-1 ℃ to form flocculent colonies, placing pearl cotton around the first culture dish, spreading rhizopus rhizoids and sporangiums and growing the rhizopus rhizoids and sporangiums on the pearl cotton, and selecting sparsely distributed rhizopus rhizoids and sporangiums from the pearl cotton to manufacture the slide specimen.
Preferably, in the method for manufacturing the rhizopus slide specimen, the first culture dish is placed on the pearl wool, and the pearl wool is ensured to be reserved around the bottom of the first culture dish.
Preferably, in the method for manufacturing the rhizopus slide specimen, when the pearl cotton is placed around the first culture dish, the first culture dish with the pearl cotton is placed in the second culture dish, the diameter and the height of the first culture dish are both smaller than those of the second culture dish, the first culture dish is not provided with a dish cover, and the second culture dish is provided with a dish cover.
More preferably, in the method for manufacturing a rhizopus slide specimen, when the pearl wool is placed around the first culture dish, the pearl wool is round, the diameter of the pearl wool is equal to the inner diameter of the second culture dish, the pearl wool is laid in the second culture dish, and then the first culture dish is placed on the upper portion of the pearl wool.
Preferably, when the pearl cotton is placed around the first culture dish, the pearl cotton is round, the diameter of the pearl cotton is equal to the inner diameter of the second culture dish, the bottom of the pearl cotton is provided with a groove or a through hole, and then the bottom of the first culture dish is embedded in the groove or the through hole.
Preferably, the method for preparing the rhizopus slide specimen comprises the following steps of:
carefully selecting sparsely distributed rhizopus from pearl cotton, placing the rhizopus on a glass slide, dripping 1 drop of alcohol with volume concentration of 80-95% to wet the rhizopus on the glass slide, adding 1 drop of potassium hydroxide solution with mass concentration of 2-3% when the alcohol is not completely volatilized, correcting the contraction action of the alcohol to restore the material, then sucking off redundant potassium hydroxide solution by using absorbent paper, adding 1 drop of liquid sealing agent, covering a cover plate, sucking off the redundant sealing agent, and manufacturing to obtain a complete rhizopus slide specimen.
More preferably, the method for selecting sparsely distributed rhizopus from pearl cotton comprises the following steps: rhizopus spread on pearl wool, which is seen as a single black spot with naked eyes around the cotton.
Preferably, the sealing agent is prepared by mixing and stirring gum arabic, double distilled water, glycerin and chloral hydrate at normal temperature, wherein the ratio of the gum arabic to the double distilled water to the glycerin to the chloral hydrate is 3g:5mL:2mL:20 g.
Preferably, the pearl cotton is pure white pearl cotton.
More preferably, the density of the pearl cotton is 0.025-0.035 g/cm3A compressive strength of 3.0 to 4.0kg/cm3The water absorption rate is 0.01-0.015 kg/cm3The recovery rate is more than 98%.
Preferably, the medium is PDA medium.
The second purpose of the invention is to provide a rhizopus slide specimen, which is prepared by the preparation method of the rhizopus slide specimen.
Compared with the prior art, the rhizopus slide specimen and the manufacturing method thereof have the following beneficial effects:
in order to clearly observe sporangium and rhizoid, the step of selecting rhizopus is found to be the main factor causing incomplete rhizopus in routine experiments, the invention further contemplates starting from the material for growth and growth of rhizopus and exploring the material which is easier to select and does not damage the complete structure of rhizopus. The pearl cotton has excellent rhizopus colonization growth conditions and easy attachment, is suitable for creeping and spreading of rhizopus, and has the characteristics of compression elasticity, recovery rate and water absorption rate, especially the characteristics of suitability for picking rhizopus, which can not damage the integrity of rhizopus, so that the prepared rhizopus slide specimen can clearly see the main identification characteristics of sporangium and rhizoid, observe the form of rhizopus and has good effect.
Drawings
FIG. 1 is a schematic view showing the hypha extension of Rhizopus in example 2 of the present invention colliding with pearl wool and growing;
FIG. 2 is a vertical cross-sectional view of the culture dish of FIG. 1.
Description of reference numerals: 1. a first culture dish; 2. a second culture dish; 3. pearl wool; 4. a culture medium; 5. flocculent colonies of rhizopus; 6. the rhizopus which touches pearl cotton and grows in a growing way is spread.
Detailed Description
A specific embodiment of the present invention is described in detail below, but it should be understood that the scope of the present invention is not limited by the specific embodiment.
Example 1
The invention aims to provide a method for manufacturing a rhizopus slide specimen, which comprises the steps of placing a culture medium at the bottom of a first culture dish, inoculating rhizopus in the center of the bottom of the first culture dish, culturing at 25 +/-1 ℃ to form flocculent colonies, placing pearl cotton around the first culture dish, allowing rhizopus rhizoids and sporangiums to spread and grow on the pearl cotton, and selecting sparsely distributed rhizopus rhizoids and sporangiums from the pearl cotton to manufacture the slide specimen.
Wherein the culture medium is a PDA culture medium.
When specifically placing the pearl cotton around first culture dish, place first culture dish on the pearl is cotton, and guarantee to remain the pearl cotton around first culture dish bottom, will place the whole placing on clean filter paper of first culture dish on the pearl cotton.
The method for preparing the rhizopus slide specimen comprises the following steps of selecting sparsely distributed rhizopus rhizoids and sporangium from pearl cotton to prepare the slide specimen: the method comprises the steps of carefully placing the sparsely distributed rhizopus on a glass slide from pearl cotton, dripping 1 drop of alcohol with volume concentration of 80% to wet the rhizopus on the glass slide, adding 1 drop of potassium hydroxide solution with mass concentration of 3% when the alcohol is not completely volatilized, correcting the contraction action of the alcohol to restore the material, then sucking off redundant potassium hydroxide solution by using absorbent paper, adding 1 drop of liquid sealing agent, covering a cover plate, sucking off the redundant sealing agent, and thus obtaining a complete rhizopus slide specimen.
More preferably, the method for selecting sparsely distributed rhizopus from pearl cotton comprises the following steps: rhizopus spread on pearl wool, which is seen as a single black spot with naked eyes around the cotton.
In this embodiment, the sealant is prepared by mixing and stirring gum arabic, double distilled water, glycerin, and chloral hydrate at room temperature, and the formula of the sealant is as follows: 30g of Arabic gum, 200g of chloral hydrate, 20mL of glycerol and 50mL of distilled water; the preparation method comprises the following steps: placing gum arabic into a beaker, slowly adding distilled water and stirring, heating in water bath, slowly adding mixed solution of chloral hydrate while stirring, adding glycerol, stirring, pouring into a narrow-neck bottle, covering with a glass plug, placing in a 60 deg.C incubator for 24 hr, filtering with clean glass wool and gauze in the incubator, and storing in dark place.
Wherein the pearl cotton is pure white pearl cotton with density of 0.025g/cm3Compressive strength of 3.0kg/cm3Water absorption of 0.015kg/cm3The recovery rate is more than 98%.
Example 2
As shown in fig. 1, a schematic diagram of the rhizopus in example 1 that hyphae overgrow and touch pearl cotton and grow in an overgrowth mode is shown, the diameter and height of a first culture dish 1 are both smaller than those of a second culture dish 2, specifically, in this example, the diameter and height of the first culture dish 1 are 8cm and 12cm, the diameter and height of the second culture dish 2 are 15cm and 24cm, the method for manufacturing a rhizopus slide specimen in this example is that a culture medium 4 is placed at the bottom of the first culture dish 1, rhizopus is inoculated at the center of the bottom of the first culture dish 1, then the first culture dish 1 is placed at the center of the second culture dish 2 with pearl cotton 3 laid at the bottom, specifically, when pearl cotton is placed around the first culture dish, the first culture dish 1 with pearl cotton 3 is placed in the second culture dish 2, and the first culture dish 1 is not provided with a dish cover, the second culture dish 2 is provided with a dish cover, specifically, and the selected pearl cotton is in this example is round, the diameter of the pearl wool is equal to the inner diameter of the second culture dish, the pearl wool is paved in the second culture dish, then the first culture dish is placed at the upper part of the pearl wool, then the pearl wool is cultured under the condition of 25 +/-1 ℃, firstly, rhizopus flocculent colonies 5 are formed in the first culture dish 1, as the height of the first culture dish 1 is lower than that of the second culture dish 2, the rhizopus flocculent colonies 5 cross the culture dish 1 with lower height to spread to the pearl wool 3 and grow in situ, complete rhizopus individuals grow on the pearl wool 3, 6 are rhizopus touching the pearl wool and growing in situ for spreading, and then, rhizopus pseudoroots and sporangium which are sparsely distributed are selected from the pearl wool to prepare a slide specimen.
Wherein the culture medium adopts PDA culture medium. Specifically, the method for selecting sparsely distributed rhizopus pseudoroots and sporangium from pearl cotton to prepare the slide specimen comprises the following steps:
the method comprises the steps of carefully placing the sparsely distributed rhizopus on a glass slide from pearl cotton, dripping 1 drop of alcohol with volume concentration of 90% to moisten the rhizopus on the glass slide, adding 1 drop of potassium hydroxide solution with mass concentration of 2% when the alcohol is not completely volatilized, correcting the contraction action of the alcohol to restore the material, then sucking off redundant potassium hydroxide solution by using water absorption paper, adding 1 drop of liquid sealing agent, covering a cover plate, sucking off the redundant sealing agent, and thus obtaining a complete rhizopus slide specimen.
Specifically, the method for selecting sparsely distributed rhizopus from pearl cotton comprises the following steps: rhizopus spread on pearl wool, which is seen as a single black spot with naked eyes around the cotton.
The sealing agent of this example was prepared by mixing and stirring gum arabic, double distilled water, glycerin, chloral hydrate at room temperature, and the specific preparation method was the same as in the examples.
In the embodiment, the pearl cotton is pure white pearl cotton with a density of 0.035g/cm3Compressive strength of 4.0kg/cm3Water absorption of 0.015kg/cm3The recovery rate is more than 98%.
Example 3
The method for manufacturing the rhizopus slide specimen is the same as that in embodiment 2, and the only difference is that when the pearl cotton is placed around the first culture dish, the pearl cotton is round, the diameter of the pearl cotton is equal to the inner diameter of the second culture dish, a groove or a through hole is formed in the bottom of the pearl cotton, the pearl cotton is laid in the second culture dish, and then the bottom of the first culture dish is embedded in the groove or the through hole.
Preferred embodiments of the present invention have been described for the purpose of preventing redundancy, and although preferred embodiments of the present invention have been described, additional variations and modifications in those embodiments may occur to those skilled in the art once they learn of the basic inventive concepts. Therefore, the present invention is not limited to the above-mentioned embodiments, and those skilled in the art can make various changes without inventive efforts from the above-mentioned conception, and fall within the scope of the present invention.
Claims (10)
1. A method for manufacturing a rhizopus slide specimen is characterized in that a culture medium is placed at the bottom of a first culture dish, rhizopus is inoculated at the bottom of the first culture dish, then the culture is carried out at 25 +/-1 ℃ to form flocculent colonies, pearl cotton is placed around the first culture dish, rhizopus rhizomes and sporangium of the rhizopus spread and grow on the pearl cotton, and then the sparse rhizopus containing the rhizopus rhizomes and sporangium is selected from the pearl cotton to manufacture the slide specimen.
2. The method for preparing a rhizopus slide specimen according to claim 1, wherein when the pearl wool is placed around the first culture dish, the first culture dish is placed on the pearl wool, and the pearl wool is ensured to be remained around the bottom of the first culture dish.
3. The method for preparing a rhizopus slide specimen according to claim 1, wherein when the pearl wool is placed around the first culture dish, the first culture dish with the pearl wool is placed in the second culture dish, the diameter and the height of the first culture dish are both smaller than those of the second culture dish, the first culture dish is not provided with a dish cover, and the second culture dish is provided with a dish cover.
4. The method for preparing a rhizopus slide specimen according to claim 3, wherein the pearl wool is round when the pearl wool is placed around the first culture dish, the diameter of the pearl wool is equal to the inner diameter of the second culture dish, the pearl wool is laid in the second culture dish, and then the first culture dish is placed on the upper portion of the pearl wool.
5. The method for preparing a rhizopus slide specimen according to claim 3, wherein when the pearl wool is placed around the first culture dish, the pearl wool is round, the diameter of the pearl wool is equal to the inner diameter of the second culture dish, a groove or a through hole is formed in the bottom of the pearl wool, the pearl wool is laid in the second culture dish, and then the bottom of the first culture dish is embedded in the groove or the through hole.
6. The method for preparing a rhizopus slide specimen as claimed in claim 1, wherein the method for selecting sparsely distributed rhizopus containing rhizopus pseudorhizomes and sporangiums from pearl cotton to prepare the slide specimen comprises the following steps:
carefully selecting sparsely distributed rhizopus from pearl cotton, placing the rhizopus on a glass slide, dripping 1 drop of alcohol with volume concentration of 80-95% to wet the rhizopus on the glass slide, adding 1 drop of potassium hydroxide solution with mass concentration of 2-3% when the alcohol is not completely volatilized, correcting the contraction action of the alcohol to restore the material, then sucking off redundant potassium hydroxide solution by using absorbent paper, adding 1 drop of liquid sealing agent, covering a cover plate, sucking off the redundant sealing agent, and manufacturing to obtain a complete rhizopus slide specimen.
7. The method for preparing a rhizopus slide specimen as claimed in claim 6, wherein the method for selecting sparsely distributed rhizopus from pearl wool comprises: rhizopus spread on pearl wool, which is seen as a single black spot with naked eyes around the cotton.
8. The method for preparing a rhizopus slide specimen according to claim 6, wherein the sealing agent is prepared by mixing and stirring gum arabic, double distilled water, glycerin and chloral hydrate at room temperature, wherein the ratio of gum arabic to double distilled water to glycerin to chloral hydrate is 3g:5mL:2mL:20 g.
9. The method for preparing a rhizopus slide specimen according to any one of claims 1 to 8, wherein the pearl wool is pure white pearl wool, and the density of the pearl wool is 0.025 to 0.035g/cm3A compressive strength of 3.0 to 4.0kg/cm3The water absorption rate is 0.01-0.015 kg/cm3The recovery rate is more than 98%.
10. A rhizopus slide specimen prepared by the method for preparing a rhizopus slide specimen according to any one of claims 1 to 8.
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