CN109490268A - A kind of fluorescence microscopy method of efficient, high-definition observation of plant pollen tube growing state in gynoecium - Google Patents
A kind of fluorescence microscopy method of efficient, high-definition observation of plant pollen tube growing state in gynoecium Download PDFInfo
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- CN109490268A CN109490268A CN201811447534.3A CN201811447534A CN109490268A CN 109490268 A CN109490268 A CN 109490268A CN 201811447534 A CN201811447534 A CN 201811447534A CN 109490268 A CN109490268 A CN 109490268A
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- 238000000034 method Methods 0.000 title claims abstract description 51
- 238000000799 fluorescence microscopy Methods 0.000 title claims abstract description 41
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- GEHJYWRUCIMESM-UHFFFAOYSA-L sodium sulfite Chemical compound [Na+].[Na+].[O-]S([O-])=O GEHJYWRUCIMESM-UHFFFAOYSA-L 0.000 claims description 18
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- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 16
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- 239000001045 blue dye Substances 0.000 claims description 15
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- 229940101006 anhydrous sodium sulfite Drugs 0.000 claims description 10
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- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 claims description 8
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- LRBQNJMCXXYXIU-PPKXGCFTSA-N Chinese gallotannin Chemical compound OC1=C(O)C(O)=CC(C(=O)OC=2C(=C(O)C=C(C=2)C(=O)OC[C@@H]2[C@H]([C@H](OC(=O)C=3C=C(OC(=O)C=4C=C(O)C(O)=C(O)C=4)C(O)=C(O)C=3)[C@@H](OC(=O)C=3C=C(OC(=O)C=4C=C(O)C(O)=C(O)C=4)C(O)=C(O)C=3)[C@H](OC(=O)C=3C=C(OC(=O)C=4C=C(O)C(O)=C(O)C=4)C(O)=C(O)C=3)O2)OC(=O)C=2C=C(OC(=O)C=3C=C(O)C(O)=C(O)C=3)C(O)=C(O)C=2)O)=C1 LRBQNJMCXXYXIU-PPKXGCFTSA-N 0.000 claims description 4
- 239000000975 dye Substances 0.000 claims description 3
- 240000002834 Paulownia tomentosa Species 0.000 claims 1
- 238000004043 dyeing Methods 0.000 abstract description 10
- 230000000694 effects Effects 0.000 abstract description 10
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 abstract description 9
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- 238000003199 nucleic acid amplification method Methods 0.000 abstract description 5
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- 238000002474 experimental method Methods 0.000 abstract description 2
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- 238000007796 conventional method Methods 0.000 description 12
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- 238000002360 preparation method Methods 0.000 description 5
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- 229930002875 chlorophyll Natural products 0.000 description 4
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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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- 239000000203 mixture Substances 0.000 description 3
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- 229920000018 Callose Polymers 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 2
- 238000005286 illumination Methods 0.000 description 2
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
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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
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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
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N2021/6439—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" with indicators, stains, dyes, tags, labels, marks
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- Health & Medical Sciences (AREA)
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Abstract
The invention discloses a kind of fluorescence microscopy methods of observation of plant pollen tube growing state in gynoecium efficient, high-definition.Fluorescence microscopy method of the invention greatly weakens the luminous phenomenon of gynoecium autologous tissue, reduce its interference in larger amplification factor to pollen tube observation, it is ideal to the presentation effect of details to be easy to obtain the image high-definition of pollen tube tumor growth under high-amplification-factor.Fluorescence microscopy method of the invention greatly foreshortens to several hours for clearing time is softened, and the time of dyeing also foreshortens to a few minutes from a few hours of the prior art, so that observation experiment can be completed in 1 day, conventional efficient has been significantly increased, the NaOH solution using high concentration is avoided simultaneously, material size is able to maintain reset condition in softening clearing process, therefore without carrying out rehydration processing the phenomenon that be not in dehydration shrinkage.
Description
Technical field:
The invention belongs to microscopic examination technique fields, and in particular to a kind of observation of plant pollen tube efficient, high-definition
The fluorescence microscopy method of growing state in gynoecium.
Background technique:
Growing state of the pollen tube in gynoecium generally utilizes fluorescence microscopy observation to observe.Distinctive callosity in pollen tube
Matter inspires bright blue-green fluorescent in conjunction with the fluorchrome in colourless aniline blue under ultraviolet light, and its in gynoecium
Hetero-organization be free of or less containing callose to not shine or weak light.By this technology, pollen tube can be grown in gynoecium
Situation is effectively observed, and is the necessary means for studying the important physiology course such as plant self-compatibility and fertilization process.
Traditional pollen tube fluorescence microscopy, after transparent to the simply softening of vegetable material (such as style and ovary), warp
It is added dropwise after azure dye dyeing after aniline blue dye liquor direct tablet compressing in fluorescence microscopy under the microscope.In conventional method, general benefit
Carry out immersion softening with NaOH solution, required time is long, is greater than 1 millimeter of sturdy material especially for style diameter, it is few then one
Then more than ten days more than two days, and prolonged impregnate may not necessarily also obtain ideal observing effect.Also someone uses before softening
NaClO carries out transparent processing, but regardless of which kind of mode not can avoid certain vascular tissues after this kind of conventional process
The spontaneous hyperfluorescence of autologous tissue in flourishing sturdy style or ovary, thus the observation to pollen tube is interfered, some even nothings
Method differentiates pollen tube and style tissue.Therefore observing effect is not good enough, is difficult to be accurate to the details of single pollen tube.
Summary of the invention:
The purpose of the present invention is overcoming defect in the prior art, a kind of efficient, high-definition observation of plant flower is provided
The fluorescence microscopy method of tube cell growing state in gynoecium.
The fluorescence microscopy method of observation of plant pollen tube growing state in gynoecium of the invention, comprising the following steps:
(1) it using the gynoecium after plant to be seen pollination as material to be checked, is immersed in FAA fixer and fixes 4h or more；
This step is to fix materials microstructure to be checked, and eliminate material Determination of Chlorophyll to be checked in case its in subsequent Fluirescence observation from
Sending out red fluorescence influences observing effect.
(2) by the material to be checked after fixation wash with distilled water after, material to be checked is placed in container, to equipped with to sample
The anhydrous sodium sulfite solution of mass fraction 10% is added in the container of material, dosage, which is subject to, all submerges material to be checked, then
Container is placed in 90-100 DEG C of water-bath after the closely translucent g., jelly-like of material softening to be checked to yellow to take out；Softening process
In, solution can become yellowish-brown gradually；Depending on material size, the time can reach perfect condition in 2-3h or so.This step is
Soften transparent material, observed convenient for subsequent tabletting, and weakens tissue autofluorescence phenomenon.
(4) the anhydrous sodium sulfite solution in container is siphoned away, after being washed with distilled water, the dyeing of decolorized aniline blue dye liquor is added
1-5 minutes；Soften transparent good material, dyeing is rapid, a few minutes.Too long dyeing time can make instead pollen tube with
Outer tissue catches pigment to also fluoresce when Fluirescence observation, influences the observing effect to pollen tube.
(5) material to be checked dyed taking-up is placed on glass slide, coverslip tabletting is used after glycerite is added dropwise, in purple
Fluorescence microscopy observation is carried out under outer light.Glycerol mounting can increase the transparency of microscopic observation, guarantee observation when larger amplification factor
Clarity, and piece save when will not dry out because of dehydration.
It is preferred that the FAA fixer includes the ethanol water, glacial acetic acid and formaldehyde of volume fraction 50%, it is described
The volume ratio of the ethanol water of volume fraction 50%, glacial acetic acid and formaldehyde is 18:1:1.
The decolorized aniline blue dye liquor is preferably the colourless aniline blue dye liquor of mass fraction 0.1%.
The glycerite is preferably the glycerine water solution of mass fraction 75%.
The regular time is preferably 4~8h.
The plant to be seen is preferably seashore paulownia, tomonea, Radix Mussaendae or Lisianthus.
The container is preferably 2.5mL centrifuge tube.
Compared with prior art, the invention has the following advantages:
Firstly, this method processing is lower to plant present invention employs the material softening clearing method completely different with the prior art
Object gynoecium autologous tissue's luminescence phenomenon is greatly weakened, and is reduced it and is done in larger amplification factor to pollen tube observation
It disturbs, to be easy to obtain the image high-definition grown in pollen tube gynoecium under high-amplification-factor, to the presentation effect of details
It is ideal.Secondly, the present invention, which will soften clearing time, greatly foreshortens to several hours, and the time dyed is also from existing skill
The a few hours of art foreshorten to a few minutes, so that observation experiment can be completed in 1 day, conventional efficient has been significantly increased.Furthermore
Fluorescence microscopy method of the invention avoids the NaOH solution using high concentration, softens material size in clearing process and is able to maintain original
Beginning state, therefore without carrying out rehydration processing the phenomenon that be not in dehydration shrinkage.In addition, in mounting observation, the prior art
Using aniline blue dye liquor mounting, but the present invention uses glycerol mounting, because the glycerine water solution of mass fraction 75% has and glass
Similar refractive index significantly improves transparency when observation, so that micro-imaging effect is further improved.
Detailed description of the invention:
Fig. 1 is pollen tube growth situation (× 100 in seashore paulownia pistil tissue when embodiment 1 does not use FAA fixer fixed
Times)；Wherein A is the chlorophyll of spontaneous red fluorescence.
Fig. 2 be embodiment 2 seashore paulownia pollination after column cap and style in pollen tube growth situation (× 40 times).
Fig. 3 be embodiment 2 the pollination of seashore paulownia after pollen tube growth situation (× 40 times) in column cap part；Wherein A is to pass
The processing of system method；B is that the fluorescence microscopy method of the present embodiment is handled.
Fig. 4 is the effect that pollen tube is observed under the high-amplification-factor of column cap part after the seashore paulownia of embodiment 2 pollinates；Wherein 1&
2 be 100 times of amplification, and 3&4 is 200 times of amplification.
Fig. 5 be embodiment 3 tomonea pollination after pollen tube growth situation in column cap part；Wherein A-1 is conventional method
Handle (× 100 times), A-2 be conventional process (× 200 times), B-1 be the present embodiment fluorescence microscopy method processing (×
100 times), B-2 is that the fluorescence microscopy method of the present embodiment handles (× 200 times).
Fig. 6 be embodiment 4 Radix Mussaendae pollination after pollen tube growth situation in style part；Wherein A-1 is tradition side
Method handle (× 200 times), A-2 be conventional process (× 400 times), B-1 be the present embodiment fluorescence microscopy method processing (×
200 times), B-2 is that the fluorescence microscopy method of the present embodiment handles (× 400 times).
Fig. 7 be embodiment 5 Lisianthus pollination after pollen tube growth situation in style part；Wherein A-1 is conventional method
Handle (× 100 times), A-2 be conventional process (× 200 times), B-1 be the present embodiment fluorescence microscopy method processing (×
100 times), B-2 is that the fluorescence microscopy method of the present embodiment handles (× 200 times).
Fig. 8 is pollen tube growth situation in ovary part after the Lisianthus pollination of embodiment 5；Wherein A-1 is conventional method
Handle (× 100 times), A-2 be conventional process (× 200 times), B-1 be the present embodiment fluorescence microscopy method processing (×
100 times), B-2 is that the fluorescence microscopy method of the present embodiment handles (× 200 times).
Specific embodiment:
The following examples are further illustrations of the invention, rather than limiting the invention.
Embodiment 1:
Seashore paulownia flower after choosing pollination 12h, removing petal etc. only leave gynoecium part as material to be checked, will be to
Sample material is placed in 2.5mL centrifuge tube, and the anhydrous sodium sulfite solution of mass fraction 10% is added, and dosage is to be checked all to submerge
It subject to material, is taken out after centrifuge tube is then placed in 100 DEG C of water-bath 2h, material softening to be checked turns yellow nearly translucent jelly
Shape.The anhydrous sodium sulfite solution in pipe is siphoned away, after carefully washing 3 times with distilled water, the colourless benzene of mass fraction 0.1% is added
Amine indigo plant dye liquor (preparation of the colourless aniline blue dye liquor of mass fraction 0.1%: weighs 0.1g aniline blue and is dissolved in 99.9g concentration and be
The K of 0.033mol/L3PO4In aqueous solution, the aniline blue dye liquor of mass fraction 0.1% is prepared, after avoid light place is extremely decolourized,
The colourless aniline blue dye liquor of mass fraction 0.1% is obtained, is kept in dark place spare) dyeing 2 minutes.It is carefully taken out with tweezers to sample
Material is placed on glass slide, is used coverslip tabletting after the glycerine water solution of mass fraction 75% is added dropwise, is carried out fluorescence under ultraviolet light
Microexamination (Fig. 1).
From figure 1 it appears that if FAA fixer is not used to fix, it is green containing a large amount of leaf in seashore paulownia pistil tissue
Element, in Fluirescence observation, the spontaneous red fluorescence of chlorophyll influences observing effect.
Embodiment 2:
Seashore paulownia flower after choosing pollination 12h, removing petal etc. only leave gynoecium part as material to be checked, submergence
In FAA fixer, (FAA fixer is by ethanol water, glacial acetic acid and the formaldehyde of the volume fraction 50% of volume ratio 18:1:1
Mix) in fix 4h, take out it is fixed after material to be checked, 3 times wash with distilled water, by material to be checked be placed in 2.5mL from
In heart pipe, the anhydrous sodium sulfite solution of mass fraction 10% is added, dosage, which is subject to, all submerges materials to be checked, then will be from
Heart pipe takes out after being placed in 90 DEG C of water-bath 2h, and material softening to be checked turns yellow nearly translucent g., jelly-like.It siphons away anhydrous in pipe
After carefully washing 3 times with distilled water, the colourless aniline blue dye liquor (preparation method of mass fraction 0.1% is added in sodium sulfite solution
With embodiment 1) dyeing 2 minutes.Material to be checked is carefully taken out with tweezers to be placed on glass slide, and the glycerol of mass fraction 75% is added dropwise
Coverslip tabletting is used after aqueous solution, carries out fluorescence microscopy observation (Fig. 2-4) under ultraviolet light.
It shines and style from the visible only pollen tube of fluorescence microscopy method processing that can be seen that the present embodiment in Fig. 2-4
It hardly shines, also shines without chlorophyll, the imaging effect of pollen tube is very clear with column cap tissue.With conventional method
(fluorescence microscopy method disclosed in CN201310236666.2) is compared, hence it is evident that though the visible pollen tube of visible conventional method,
Style tissue illumination by force can block pollen tube very much, and the fluorescence microscopy method style tissue of the present embodiment hardly shines, and
And good dark field can be obtained under high-amplification-factor, so that the imaging of pollen tube is very clear.
Embodiment 3:
Tomonea flower after choosing pollination 12h, removing petal etc. only leave gynoecium part as material to be checked, submergence
In FAA fixer, (FAA fixer is by ethanol water, glacial acetic acid and the formaldehyde of the volume fraction 50% of volume ratio 18:1:1
Mix) in fix 8h, take out it is fixed after material to be checked, 5 times wash with distilled water, by material to be checked be placed in 2.5mL from
In heart pipe, the anhydrous sodium sulfite solution of mass fraction 10% is added, dosage, which is subject to, all submerges materials to be checked, then will be from
Heart pipe takes out after being placed in 100 DEG C of water-bath 3h, and material softening to be checked turns yellow nearly translucent g., jelly-like.It siphons away anhydrous in pipe
After carefully washing 5 times with distilled water, the colourless aniline blue dye liquor (preparation method of mass fraction 0.1% is added in sodium sulfite solution
With embodiment 1) dyeing 1 minute.Material to be checked is carefully taken out with tweezers to be placed on glass slide, and the glycerol of mass fraction 75% is added dropwise
Coverslip tabletting is used after aqueous solution, carries out fluorescence microscopy observation (Fig. 5) under ultraviolet light.
From figure 5 it can be seen that compared with conventional method (fluorescence microscopy method disclosed in CN201310236666.2),
It is handled under conventional method, pollen tube is because style and column cap tissue illumination interfere, and image blur is unintelligible, and the fluorescence of the present embodiment
It is clearly demarcated that microscopic method handles pollen tube root root.And conventional process, pistil tissue are shunk, and it is visible under same equimultiple
Tissue and pollen tube become smaller very much.
Embodiment 4:
Radix Mussaendae flower after choosing pollination 12h, removing petal etc. only leave gynoecium part as material to be checked, leaching
In FAA fixer, (FAA fixer is not by ethanol water, glacial acetic acid and the first of the volume fraction 50% of volume ratio 18:1:1
Aldehyde mixes) in fix 4h, take out it is fixed after material to be checked, 3 times wash with distilled water, material to be checked is placed in 2.5mL
In centrifuge tube, the anhydrous sodium sulfite solution of mass fraction 10% is added, dosage, which is subject to, all submerges material to be checked, then will
Centrifuge tube takes out after being placed in 100 DEG C of water-bath 2h, and material softening to be checked turns yellow nearly translucent g., jelly-like.Siphon away the nothing in pipe
After carefully washing 3 times with distilled water, colourless aniline blue dye liquor (the preparation side of mass fraction 0.1% is added in water sodium sulfite solution
Method is with embodiment 1) dyeing 5 minutes.Material to be checked is carefully taken out with tweezers to be placed on glass slide, and the sweet of mass fraction 75% is added dropwise
Coverslip tabletting is used after oil solution, carries out fluorescence microscopy observation (Fig. 6) under ultraviolet light.
From fig. 6 it can be seen that compared with conventional method (fluorescence microscopy method disclosed in CN201310236666.2),
It can be seen that under conventional method, when high magnification numbe, pollen tube is blocked because style tissue, can when taking pictures in addition to bright callose plug
See, it is virtually impossible to recognize pollen tube, and pollen tube is clear in structure under the fluorescence microscopy method of the present embodiment is handled, root root is clearly demarcated.
Embodiment 5:
Lisianthus flower after choosing pollination 12h, removing petal etc. only leave gynoecium part as material to be checked, submergence
In FAA fixer, (FAA fixer is by ethanol water, glacial acetic acid and the formaldehyde of the volume fraction 50% of volume ratio 18:1:1
Mix) in fix 4h, take out it is fixed after material to be checked, 3 times wash with distilled water, by material to be checked be placed in 2.5mL from
In heart pipe, the anhydrous sodium sulfite solution of mass fraction 10% is added, dosage, which is subject to, all submerges materials to be checked, then will be from
Heart pipe takes out after being placed in 100 DEG C of water-bath 3h, and material softening to be checked turns yellow nearly translucent g., jelly-like.It siphons away anhydrous in pipe
After carefully washing 3 times with distilled water, the colourless aniline blue dye liquor (preparation method of mass fraction 0.1% is added in sodium sulfite solution
With embodiment 1) dyeing 2 minutes.Material to be checked is carefully taken out with tweezers to be placed on glass slide, and the glycerol of mass fraction 75% is added dropwise
Coverslip tabletting is used after aqueous solution, carries out fluorescence microscopy observation (Fig. 7-8) under ultraviolet light.
It can be seen that and conventional method (fluorescence microscopy method disclosed in CN201310236666.2) phase from Fig. 7-8
Than, it is seen that under conventional method, style and ovary tissues are blocked, image blur, it is difficult to the blur-free imaging of single pollen tube is obtained, and
Image pollen tube obtained by the fluorescence microscopy method of the present embodiment is clear in structure.
Claims (6)
1. a kind of fluorescence microscopy method of observation of plant pollen tube growing state in gynoecium, which is characterized in that including following step
It is rapid:
(1) it using the gynoecium after plant to be seen pollination as material to be checked, is immersed in FAA fixer and fixes 4h or more；
(2) by the material to be checked after fixation wash with distilled water after, material to be checked is placed in container；To equipped with material to be checked
The anhydrous sodium sulfite solution of mass fraction 10% is added in container, dosage, which is subject to, all submerges material to be checked, then will hold
Device is placed in 90-100 DEG C of water-bath and takes out after the closely translucent g., jelly-like of material softening to be checked to yellow；
(3) the anhydrous sodium sulfite solution in container is siphoned away, after being washed with distilled water, decolorized aniline blue dye liquor is added and dyes 1-5
Minute；
(4) material to be checked dyed taking-up is placed on glass slide, coverslip tabletting is used after glycerite is added dropwise, in ultraviolet light
Lower progress fluorescence microscopy observation.
2. the fluorescence microscopy method of observation of plant pollen tube growing state in gynoecium according to claim 1, feature
It is, the FAA fixer includes the ethanol water, glacial acetic acid and formaldehyde of volume fraction 50%, the volume fraction
The volume ratio of 50% ethanol water, glacial acetic acid and formaldehyde is 18:1:1.
3. the fluorescence microscopy method of observation of plant pollen tube growing state in gynoecium according to claim 1, feature
It is, the decolorized aniline blue dye liquor is the colourless aniline blue dye liquor of mass fraction 0.1%.
4. the fluorescence microscopy method of observation of plant pollen tube growing state in gynoecium according to claim 1, feature
It is, the glycerite is the glycerine water solution of mass fraction 75%.
5. the fluorescence microscopy side of observation of plant pollen tube growing state in gynoecium according to any one of claims 1 to 4
Method, which is characterized in that the plant to be seen is seashore paulownia, tomonea, Radix Mussaendae or Lisianthus.
6. the fluorescence microscopy method of observation of plant pollen tube growing state in gynoecium according to claim 1, feature
It is, the container is 2.5mL centrifuge tube.
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| CN110583634A (en) * | 2019-10-13 | 2019-12-20 | 安徽省农业科学院水稻研究所 | Preservation method for length of rice stigma |
| CN110583634B (en) * | 2019-10-13 | 2022-01-14 | 安徽省农业科学院水稻研究所 | Preservation method of rice stigma |
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