CN116380595A - Simple and effective bamboo leaf epidermis flaking method - Google Patents
Simple and effective bamboo leaf epidermis flaking method Download PDFInfo
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- CN116380595A CN116380595A CN202310490367.5A CN202310490367A CN116380595A CN 116380595 A CN116380595 A CN 116380595A CN 202310490367 A CN202310490367 A CN 202310490367A CN 116380595 A CN116380595 A CN 116380595A
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- 238000000034 method Methods 0.000 title claims abstract description 41
- 235000017166 Bambusa arundinacea Nutrition 0.000 title claims abstract description 36
- 235000017491 Bambusa tulda Nutrition 0.000 title claims abstract description 36
- 241001330002 Bambuseae Species 0.000 title claims abstract description 36
- 235000015334 Phyllostachys viridis Nutrition 0.000 title claims abstract description 36
- 239000011425 bamboo Substances 0.000 title claims abstract description 36
- 210000002615 epidermis Anatomy 0.000 title claims abstract description 29
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 42
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- 239000012153 distilled water Substances 0.000 claims abstract description 14
- 238000004043 dyeing Methods 0.000 claims abstract description 14
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- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 claims abstract description 9
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- AWJWCTOOIBYHON-UHFFFAOYSA-N furo[3,4-b]pyrazine-5,7-dione Chemical compound C1=CN=C2C(=O)OC(=O)C2=N1 AWJWCTOOIBYHON-UHFFFAOYSA-N 0.000 claims abstract description 9
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- 238000005204 segregation Methods 0.000 claims abstract description 8
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- 239000006059 cover glass Substances 0.000 claims description 17
- OARRHUQTFTUEOS-UHFFFAOYSA-N safranin Chemical compound [Cl-].C=12C=C(N)C(C)=CC2=NC2=CC(C)=C(N)C=C2[N+]=1C1=CC=CC=C1 OARRHUQTFTUEOS-UHFFFAOYSA-N 0.000 claims description 12
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- 230000000694 effects Effects 0.000 abstract description 11
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- 239000000463 material Substances 0.000 description 8
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- ATNHDLDRLWWWCB-AENOIHSZSA-M chlorophyll a Chemical compound C1([C@@H](C(=O)OC)C(=O)C2=C3C)=C2N2C3=CC(C(CC)=C3C)=[N+]4C3=CC3=C(C=C)C(C)=C5N3[Mg-2]42[N+]2=C1[C@@H](CCC(=O)OC\C=C(/C)CCC[C@H](C)CCC[C@H](C)CCCC(C)C)[C@H](C)C2=C5 ATNHDLDRLWWWCB-AENOIHSZSA-M 0.000 description 4
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- 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
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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
- G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
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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
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
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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
- G01N1/34—Purifying; Cleaning
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- G01N1/00—Sampling; Preparing specimens for investigation
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- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/44—Sample treatment involving radiation, e.g. heat
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- G01N21/84—Systems specially adapted for particular applications
- G01N2021/8466—Investigation of vegetal material, e.g. leaves, plants, fruits
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Abstract
The invention discloses a simple and effective bamboo leaf epidermis flaking method, which comprises the following steps: blade collection: taking fresh green bamboo mature leaves, taking the middle positions of the leaves, and cutting the leaves into a plurality of small pieces, wherein the length of each small piece of leaves is about 0.8cm-1.2cm, and the width of each small piece of leaves is about 0.6cm-0.8cm; blade treatment: immersing the small leaf into chromic acid: nitric acid = 1:1, then placing the solution into a baking oven at 45 ℃ for segregation, and periodically replacing the saturated liquid during the segregation until the blade becomes a film-like shape and floats on the solution; cleaning: picking up the film-shaped blade by forceps for cleaning; dyeing: dyeing the cleaned blade with a dye liquor for 10min, and then washing the blade with distilled water to remove the dye liquor; and (3) tabletting: preparing a temporary loading piece by using a glass slide; and (3) observing and photographing: the simple and effective bamboo leaf epidermis flaking method is simple to operate, short in period, low in cost and good in observation effect.
Description
Technical Field
The invention relates to the technical field of plant microscopic observation, in particular to a microscopic flaking method for observing and researching the epidermis of a bamboo leaf by treating the bamboo leaf.
Background
Leaf blades are one of important organs of plants, are main organs for photosynthesis, transpiration and synthesis of organic substances, and have important influence on plant growth due to structural characteristics. The plant leaf epidermis preparation is an important means for researching the plant leaf epidermis character, and has wide application in the aspects of taxonomy, systematics, plant physiology and the like. Common methods for rapid plant leaf epidermis production include epidermis tearing, scotch tape sticking, nail polish blotting, dissociation, scraping, etc., but different methods are applicable to different plants. The bamboo plant leaf matter is thin, and silicon cells in the epidermal cells make bamboo She Cucao, so that many methods for quickly observing the plant leaf epidermis are not applicable, such as an epidermis tearing method, a transparent adhesive tape sticking method and the like, and a dissociation method utilizes chemical reagents to dissolve middle layer substances among cells to separate the cells.
Disclosure of Invention
The invention aims to overcome the existing defects, provides a simple, convenient and effective bamboo leaf epidermis flaking method, has the advantages of simple operation, short period, low cost and good observation effect, and can effectively solve the problems in the background technology.
In order to achieve the above purpose, the present invention provides the following technical solutions: a simple and effective bamboo leaf epidermis flaking method comprises the following steps:
blade collection: taking fresh green bamboo mature leaves, taking the middle positions of the leaves, and cutting the leaves into a plurality of small blocks;
blade treatment: immersing the small leaf into chromic acid: nitric acid = 1:1, then placing the solution into a baking oven at 45 ℃ for segregation, and periodically replacing the saturated liquid during the segregation until the blade becomes a film-like shape and floats on the solution;
cleaning: picking up the film-shaped blade by forceps for cleaning;
dyeing: dyeing the cleaned blade with a dye liquor for 10min, and then washing the blade with distilled water to remove the dye liquor;
and (3) tabletting: preparing a temporary loading piece by using a glass slide;
and (3) observing and photographing: the method is simple to operate, short in period, low in cost and good in observation effect.
Further, the length of the small blade is about 0.8cm-1.2cm, and the width of the small blade is about 0.6cm-0.8cm, so that the burr structure at the blade edge of the blade can be observed completely.
Further, the concentration of the chromic acid is 10%, the concentration of the nitric acid is 10%, coconut meat of the leaves is removed, and the materials such as veins of the leaves are fixed.
Further, the time length of the segregation is 36-72 h, and the segregated blade is not only suitable for observing the air holes of the blade, but also suitable for observing morphological characteristics of She Maimai sequence, epidermis structure and the like.
Further, the thin film-shaped blades are picked up by forceps and are repeatedly washed by distilled water for 3 times, and then are immersed in 70% alcohol for secondary washing, so that chlorophyll in the blades is removed, and the observation effect is prevented from being influenced.
Furthermore, the dye liquor is 1% of safranin, the safranin has high dyeing brightness, obvious color, more striking appearance and convenient observation.
Further, during the preparation, wipe slide glass and coverslip, spread the blade after dyeing on the water droplet of slide glass completely, put down flat coverslip and use the thumb to press gently, wipe off the drop of water at edge, improve observation effect.
Furthermore, the microscope is an LEICADM2500 microscope, and an angle-adjustable eye lens barrel can be optionally matched, so that the fatigue of an operator is greatly reduced, the light flux is high, and the resolution and the depth of field are higher.
Compared with the prior art, the invention has the beneficial effects that: the simple and effective bamboo leaf epidermis flaking method has the following advantages:
compared with the prior art, the method cuts the green bamboo leaves into small pieces with the length of about 0.8cm-1.2cm and the width of about 0.6cm-0.8cm, is convenient for completely placing the small pieces under the lens of a microscope, is convenient for completely observing the leaf edge burr structure of the leaves, and immerses the small pieces in 10% chromic acid: 10% nitric acid = 1: in the solution prepared by the method 1, coconut meat of the leaf is removed, and materials such as veins of the leaf are fixed, so that air holes of the leaf are conveniently observed, and meanwhile, she Maimai sequences and morphological characteristics such as a epidermis structure are observed.
Drawings
FIG. 1 is a flow chart of a bamboo leaf epidermis flaking method of the present invention;
FIG. 2 shows the structure of the upper epidermis leaf edge of the green bamboo of the present invention;
FIG. 3 shows the structures of the epidermal alveolar cells, veins and the like of the green bamboo;
FIG. 4 is an enlarged view showing the structure of the epidermal alveolar cells and veins of the green bamboo according to the present invention;
FIG. 5 shows the structure of epidermal cells at the main and collateral vessels of the lower epidermis of the green bamboo;
FIG. 6 shows the structure of epidermal cells at the main vein of the lower epidermis of the green bamboo according to the present invention;
FIG. 7 shows the air holes and lateral vein layout of the lower epidermis of the green bamboo according to the present invention;
FIG. 8 is a view showing the structure of the pores, the coat, etc. of the lower epidermis of the green bamboo according to the present invention.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Referring to fig. 1-8, the present embodiment provides a technical solution:
embodiment one: a simple and effective bamboo leaf epidermis flaking method comprises the following steps:
blade collection: fresh green bamboo mature leaves are taken, cleaned by clean water, water on the leaves is wiped off by water absorbing paper, so that the leaves are kept clean, tidy and flat, a leaf handle is sheared, 1/3 part of the middle of each leaf is reserved, a plurality of small blocks with the length of about 0.8cm and the width of about 0.6cm are sheared, the force is uniform when the leaves are sheared, the breakage of tissues is avoided, the small blocks of leaves are conveniently and completely placed under a lens of a microscope, and the leaf edge burr structure of each leaf is conveniently and completely observed.
Blade treatment: immersing the small pieces of leaf into 10% chromic acid: 10% nitric acid = 1: in the solution prepared by the method 1, the blade is completely immersed in the solution, the dosage of the fixing solution is more than 20 times of the volume of the blade, the concentration of the fixing solution is prevented from being diluted by water in the blade, so that the fixing effect of the blade material is affected, the blade is placed into a 45 ℃ oven for isolation for 36 hours, the soaking solution is replaced at regular time until the blade becomes a film-shaped floating on the solution, the coconut meat of the blade is removed, the air holes of the blade are conveniently observed, and meanwhile, the morphology characteristics such as She Maimai sequence and the epidermis structure are observed.
Cleaning: the thin film-shaped blades are lifted by forceps and are repeatedly washed for 3 times by distilled water, so that the distilled water permeates into the blades, the saturated solution is thoroughly replaced, and then the blades are immersed in 70% alcohol for secondary washing, so that chlorophyll in the blades is removed, and the observation effect is prevented from being influenced.
Dyeing: the safranin is fully filtered, then diluted to 1% concentration, the washed blade is dyed with 1% safranin for 10min, then distilled water is used for washing out the dye liquor, the safranin dyeing has high brightness, obvious color and luster, and is more striking in appearance and convenient to observe.
And (3) tabletting: the slide glass and the cover glass are wiped, water drops are arranged in the center of the slide glass by a dropper, the dyed blades are placed on the water drops of the slide glass by forceps, the surfaces of the blades are paved flat and are fully unfolded, then the cover glass is clamped by the forceps, the edge of the cover glass is contacted with the edge of the water drops on the left side of the blade material, the flat cover glass is placed downwards, the thumb is used for lightly pressing the cover glass, and the water drops on the edge are wiped by absorbent paper.
And (3) observing and photographing: and photographing under a microscope to observe structures such as leaf edge thorns, follicular cells, veins, pores, epidermal hairs and the like of the leaf.
Embodiment two:
the difference between this embodiment and the first embodiment is that:
in this embodiment, a simple and effective bamboo leaf epidermis flaking method includes the following steps:
blade collection: fresh green bamboo mature leaves are taken, cleaned by clean water, water on the leaves is wiped off by water absorbing paper, so that the leaves are kept clean, tidy and flat, a leaf handle is sheared, 1/3 part of the middle of each leaf is reserved, a plurality of small blocks with the length of about 1cm and the width of about 0.7cm are sheared, the force is uniform when the leaves are sheared, the breakage of tissues is avoided, the small blocks of leaves are conveniently and completely placed under a lens of a microscope, and the leaf edge burr structure of each leaf is conveniently and completely observed.
Blade treatment: immersing the small pieces of leaf into 10% chromic acid: 10% nitric acid = 1: in the solution prepared by the method 1, the blade is completely immersed in the solution, the dosage of the fixing solution is more than 20 times of the volume of the blade, the concentration of the fixing solution is prevented from being diluted by water in the blade, so that the fixing effect of the blade material is affected, the blade is placed into a 45 ℃ oven for isolation for 54 hours, the soaking solution is replaced at regular time until the blade becomes a film-shaped floating on the solution, the coconut meat of the blade is removed, the air holes of the blade are conveniently observed, and meanwhile, the morphology characteristics such as She Maimai sequence and the epidermis structure are observed.
Cleaning: the thin film-shaped blades are lifted by forceps and are repeatedly washed for 3 times by distilled water, so that the distilled water permeates into the blades, the saturated solution is thoroughly replaced, and then the blades are immersed in 70% alcohol for secondary washing, so that chlorophyll in the blades is removed, and the observation effect is prevented from being influenced.
Dyeing: the safranin is fully filtered, then diluted to 1% concentration, the washed blade is dyed with 1% safranin for 10min, then distilled water is used for washing out the dye liquor, the safranin dyeing has high brightness, obvious color and luster, and is more striking in appearance and convenient to observe.
And (3) tabletting: the slide glass and the cover glass are wiped, water drops are arranged in the center of the slide glass by a dropper, the dyed blades are placed on the water drops of the slide glass by forceps, the surfaces of the blades are paved flat and are fully unfolded, then the cover glass is clamped by the forceps, the edge of the cover glass is contacted with the edge of the water drops on the left side of the blade material, the flat cover glass is placed downwards, the thumb is used for lightly pressing the cover glass, and the water drops on the edge are wiped by absorbent paper.
And (3) observing and photographing: and photographing under a microscope to observe structures such as leaf edge thorns, follicular cells, veins, pores, epidermal hairs and the like of the leaf.
Embodiment III:
the difference between this embodiment and the first embodiment is that:
in this embodiment, a simple and effective bamboo leaf epidermis flaking method includes the following steps:
blade collection: fresh green bamboo mature leaves are taken, cleaned by clean water, water on the leaves is wiped off by water absorbing paper, so that the leaves are kept clean, tidy and flat, a leaf handle is sheared, 1/3 part of the middle of each leaf is reserved, a plurality of small blocks with the length of about 1.2cm and the width of about 0.8cm are sheared, the force is uniform when the leaves are sheared, the breakage of tissues is avoided, the small blocks of leaves are conveniently and completely placed under a lens of a microscope, and the leaf edge burr structure of each leaf is conveniently and completely observed.
Blade treatment: immersing the small pieces of leaf into 10% chromic acid: 10% nitric acid = 1: in the solution prepared by the method 1, the blade is completely immersed in the solution, the dosage of the fixing solution is more than 20 times of the volume of the blade, the concentration of the fixing solution is prevented from being diluted by water in the blade, so that the fixing effect of the blade material is affected, the blade is placed into a 45 ℃ oven for segregation for 72 hours, the soaking solution is replaced at regular time until the blade becomes a film-shaped floating on the solution, the coconut meat of the blade is removed, the air holes of the blade are conveniently observed, and meanwhile, the morphology characteristics such as She Maimai sequence and the epidermis structure are observed.
Cleaning: the thin film-shaped blades are lifted by forceps and are repeatedly washed for 3 times by distilled water, so that the distilled water permeates into the blades, the saturated solution is thoroughly replaced, and then the blades are immersed in 70% alcohol for secondary washing, so that chlorophyll in the blades is removed, and the observation effect is prevented from being influenced.
Dyeing: the safranin is fully filtered, then diluted to 1% concentration, the washed blade is dyed with 1% safranin for 10min, then distilled water is used for washing out the dye liquor, the safranin dyeing has high brightness, obvious color and luster, and is more striking in appearance and convenient to observe.
And (3) tabletting: the slide glass and the cover glass are wiped, water drops are arranged in the center of the slide glass by a dropper, the dyed blades are placed on the water drops of the slide glass by forceps, the surfaces of the blades are paved flat and are fully unfolded, then the cover glass is clamped by the forceps, the edge of the cover glass is contacted with the edge of the water drops on the left side of the blade material, the flat cover glass is placed downwards, the thumb is used for lightly pressing the cover glass, and the water drops on the edge are wiped by absorbent paper.
And (3) observing and photographing: and photographing under a microscope to observe structures such as leaf edge thorns, follicular cells, veins, pores, epidermal hairs and the like of the leaf.
The foregoing description is only illustrative of the present invention and is not intended to limit the scope of the invention, and all equivalent structures or equivalent processes or direct or indirect application in other related technical fields are included in the scope of the present invention.
Claims (8)
1. A simple and effective bamboo leaf epidermis flaking method is characterized in that: the method comprises the following steps:
blade collection: taking fresh green bamboo mature leaves, taking the middle positions of the leaves, and cutting the leaves into a plurality of small blocks;
blade treatment: immersing the small leaf into chromic acid: nitric acid = 1:1, then placing the solution into a baking oven at 45 ℃ for segregation, and periodically replacing the saturated liquid during the segregation until the blade becomes a film-like shape and floats on the solution;
cleaning: picking up the film-shaped blade by forceps for cleaning;
dyeing: dyeing the cleaned blade with a dye liquor for 10min, and then washing the blade with distilled water to remove the dye liquor;
and (3) tabletting: preparing a temporary loading piece by using a glass slide;
and (3) observing and photographing: and photographing and observing under a microscope.
2. The simple and effective bamboo leaf surface flaking method of claim 1, wherein the method comprises the steps of: the length of the small blade is about 0.8cm-1.2cm, and the width of the small blade is about 0.6cm-0.8cm.
3. The simple and effective bamboo leaf surface flaking method of claim 1, wherein the method comprises the steps of: the chromic acid concentration is 10%, and the nitric acid concentration is 10%.
4. The simple and effective bamboo leaf surface flaking method of claim 1, wherein the method comprises the steps of: the duration of the isolation is 36h-72h.
5. The simple and effective bamboo leaf surface flaking method of claim 1, wherein the method comprises the steps of: the thin film-like blade was picked up with forceps and rinsed back and forth 3 times with distilled water, and then immersed in 70% alcohol for a second washing.
6. The simple and effective bamboo leaf surface flaking method of claim 1, wherein the method comprises the steps of: the dye liquor is 1% safranin.
7. The simple and effective bamboo leaf surface flaking method of claim 1, wherein the method comprises the steps of: during the preparation, the slide glass and the cover glass are wiped, the dyed blade is completely unfolded on the water drop of the slide glass, the cover glass is put down, and the slide glass is lightly pressed by the thumb, so that the water drop at the edge is wiped off.
8. The simple and effective bamboo leaf surface flaking method of claim 1, wherein the method comprises the steps of: the microscope is a leicam 2500 microscope.
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| CN202310490367.5A CN116380595A (en) | 2023-05-04 | 2023-05-04 | Simple and effective bamboo leaf epidermis flaking method |
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| CN202310490367.5A CN116380595A (en) | 2023-05-04 | 2023-05-04 | Simple and effective bamboo leaf epidermis flaking method |
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| CN (1) | CN116380595A (en) |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101845415A (en) * | 2010-05-17 | 2010-09-29 | 中国科学院植物研究所 | Method for separating Casparian strip |
| CN114136738A (en) * | 2021-12-08 | 2022-03-04 | 河南中医药大学 | Microscopic flaking method for observing lower epidermis characteristics of folium artemisiae argyi |
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2023
- 2023-05-04 CN CN202310490367.5A patent/CN116380595A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101845415A (en) * | 2010-05-17 | 2010-09-29 | 中国科学院植物研究所 | Method for separating Casparian strip |
| CN114136738A (en) * | 2021-12-08 | 2022-03-04 | 河南中医药大学 | Microscopic flaking method for observing lower epidermis characteristics of folium artemisiae argyi |
Non-Patent Citations (2)
| Title |
|---|
| 张嵘梅等: "中国秋海棠属植物的叶表皮特征及其分类学意义", 云南植物研究, vol. 30, no. 06, pages 666 * |
| 魏强等: "平安竹叶片缩小的细胞学观察", 南京林业大学学报(自然科学版), vol. 43, no. 03, pages 196 * |
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