CN105486565B - A kind of method that pollen tube loads fluorescent dye - Google Patents
A kind of method that pollen tube loads fluorescent dye Download PDFInfo
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- CN105486565B CN105486565B CN201510944828.7A CN201510944828A CN105486565B CN 105486565 B CN105486565 B CN 105486565B CN 201510944828 A CN201510944828 A CN 201510944828A CN 105486565 B CN105486565 B CN 105486565B
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- pollen
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- pollen tube
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- 238000000034 method Methods 0.000 title claims abstract description 22
- 239000007850 fluorescent dye Substances 0.000 title claims abstract description 18
- 239000003086 colorant Substances 0.000 claims abstract description 20
- 239000006166 lysate Substances 0.000 claims abstract description 16
- 239000000243 solution Substances 0.000 claims abstract description 6
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims abstract description 4
- 239000000741 silica gel Substances 0.000 claims abstract description 4
- 229910002027 silica gel Inorganic materials 0.000 claims abstract description 4
- OUVXYXNWSVIOSJ-UHFFFAOYSA-N Fluo-4 Chemical group CC1=CC=C(N(CC(O)=O)CC(O)=O)C(OCCOC=2C(=CC=C(C=2)C2=C3C=C(F)C(=O)C=C3OC3=CC(O)=C(F)C=C32)N(CC(O)=O)CC(O)=O)=C1 OUVXYXNWSVIOSJ-UHFFFAOYSA-N 0.000 claims description 10
- ZCCIPPOKBCJFDN-UHFFFAOYSA-N calcium nitrate Chemical compound [Ca+2].[O-][N+]([O-])=O.[O-][N+]([O-])=O ZCCIPPOKBCJFDN-UHFFFAOYSA-N 0.000 claims description 8
- 239000002202 Polyethylene glycol Substances 0.000 claims description 6
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 6
- 229930006000 Sucrose Natural products 0.000 claims description 6
- 229920001223 polyethylene glycol Polymers 0.000 claims description 6
- 239000005720 sucrose Substances 0.000 claims description 6
- KGBXLFKZBHKPEV-UHFFFAOYSA-N boric acid Chemical compound OB(O)O KGBXLFKZBHKPEV-UHFFFAOYSA-N 0.000 claims description 4
- 239000004327 boric acid Substances 0.000 claims description 4
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 claims description 3
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 claims description 3
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 3
- HNGDOSBFYRVIEY-UHFFFAOYSA-N ethanesulfonic acid;hydrate Chemical compound O.CCS(O)(=O)=O HNGDOSBFYRVIEY-UHFFFAOYSA-N 0.000 claims description 3
- 239000007788 liquid Substances 0.000 claims 1
- 210000004027 cell Anatomy 0.000 abstract description 19
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 abstract description 13
- 229910001424 calcium ion Inorganic materials 0.000 abstract description 13
- 210000000170 cell membrane Anatomy 0.000 abstract description 4
- -1 Fluo 4AM ester Chemical class 0.000 abstract description 3
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 16
- 239000011575 calcium Substances 0.000 description 16
- 229910052791 calcium Inorganic materials 0.000 description 16
- 230000003834 intracellular effect Effects 0.000 description 10
- 230000012010 growth Effects 0.000 description 5
- 102000005701 Calcium-Binding Proteins Human genes 0.000 description 4
- 108010045403 Calcium-Binding Proteins Proteins 0.000 description 4
- 108090000371 Esterases Proteins 0.000 description 4
- 230000035515 penetration Effects 0.000 description 4
- LSTYNGLUOLXJSH-UHFFFAOYSA-N calcium boric acid dinitrate Chemical compound [N+](=O)([O-])[O-].[Ca+2].B(O)(O)O.[N+](=O)([O-])[O-] LSTYNGLUOLXJSH-UHFFFAOYSA-N 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000004744 fabric Substances 0.000 description 2
- 238000012921 fluorescence analysis Methods 0.000 description 2
- 210000003632 microfilament Anatomy 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 210000003463 organelle Anatomy 0.000 description 2
- 238000004445 quantitative analysis Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- OZLGRUXZXMRXGP-UHFFFAOYSA-N Fluo-3 Chemical compound CC1=CC=C(N(CC(O)=O)CC(O)=O)C(OCCOC=2C(=CC=C(C=2)C2=C3C=C(Cl)C(=O)C=C3OC3=CC(O)=C(Cl)C=C32)N(CC(O)=O)CC(O)=O)=C1 OZLGRUXZXMRXGP-UHFFFAOYSA-N 0.000 description 1
- 102000002151 Microfilament Proteins Human genes 0.000 description 1
- 108010040897 Microfilament Proteins Proteins 0.000 description 1
- MCEXQZRGUKALLT-VVEOGCPPSA-N acetyloxymethyl 2-[n-[2-(acetyloxymethoxy)-2-oxoethyl]-2-[2-[[6-[bis[2-(acetyloxymethoxy)-2-oxoethyl]amino]-2-[(e)-(5-oxo-2-sulfanylideneimidazolidin-4-ylidene)methyl]-1-benzofuran-5-yl]oxy]ethoxy]-4-methylanilino]acetate Chemical compound CC(=O)OCOC(=O)CN(CC(=O)OCOC(C)=O)C1=CC=C(C)C=C1OCCOC(C(=C1)N(CC(=O)OCOC(C)=O)CC(=O)OCOC(C)=O)=CC2=C1OC(\C=C\1C(NC(=S)N/1)=O)=C2 MCEXQZRGUKALLT-VVEOGCPPSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- NMUGYJRMGWBCPU-UHFFFAOYSA-N calcium orange Chemical compound C=12C=CC(=[N+](C)C)C=C2OC2=CC(N(C)C)=CC=C2C=1C(C(=C1)C([O-])=O)=CC=C1NC(=S)NC(C=1)=CC=C(N(CC(=O)OCOC(C)=O)CC(=O)OCOC(C)=O)C=1OCCOC1=CC=CC=C1N(CC(=O)OCOC(C)=O)CC(=O)OCOC(C)=O NMUGYJRMGWBCPU-UHFFFAOYSA-N 0.000 description 1
- 230000005779 cell damage Effects 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 210000000805 cytoplasm Anatomy 0.000 description 1
- 230000001086 cytosolic effect Effects 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 230000008034 disappearance Effects 0.000 description 1
- 239000000975 dye Substances 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000001215 fluorescent labelling Methods 0.000 description 1
- YFHXZQPUBCBNIP-UHFFFAOYSA-N fura-2 Chemical compound CC1=CC=C(N(CC(O)=O)CC(O)=O)C(OCCOC=2C(=CC=3OC(=CC=3C=2)C=2OC(=CN=2)C(O)=O)N(CC(O)=O)CC(O)=O)=C1 YFHXZQPUBCBNIP-UHFFFAOYSA-N 0.000 description 1
- PNDZEEPOYCVIIY-UHFFFAOYSA-N indo-1 Chemical compound CC1=CC=C(N(CC(O)=O)CC(O)=O)C(OCCOC=2C(=CC=C(C=2)C=2N=C3[CH]C(=CC=C3C=2)C(O)=O)N(CC(O)=O)CC(O)=O)=C1 PNDZEEPOYCVIIY-UHFFFAOYSA-N 0.000 description 1
- 238000000520 microinjection Methods 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000000523 sample Substances 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 210000004739 secretory vesicle Anatomy 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 208000011580 syndromic disease Diseases 0.000 description 1
- XAGUNWDMROKIFJ-UHFFFAOYSA-J tetrapotassium;2-[2-[[8-[bis(carboxylatomethyl)amino]-6-methoxyquinolin-2-yl]methoxy]-n-(carboxylatomethyl)-4-methylanilino]acetate Chemical compound [K+].[K+].[K+].[K+].C1=CC2=CC(OC)=CC(N(CC([O-])=O)CC([O-])=O)=C2N=C1COC1=CC(C)=CC=C1N(CC([O-])=O)CC([O-])=O XAGUNWDMROKIFJ-UHFFFAOYSA-J 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 230000032258 transport Effects 0.000 description 1
Classifications
-
- 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
- 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
- G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
-
- 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
- G01N2001/302—Stain compositions
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
Abstract
The invention discloses a kind of methods that pollen tube loads fluorescent dye, include the following steps：Pollen is wrapped in pan paper by the first step, is positioned in closed silica gel bottle and is stored in low temperature refrigerator；Second step after pollen takes out from refrigerator, is added in after placing 2h at normal temperatures in culture solution, it is stirred for uniformly being placed in 25 DEG C of insulating boxs and cultivates 2h, third step, by culture solution and lysate, coloring agent is corresponding mixes, 10 30min are protected from light, is subsequently placed in 25 DEG C of insulating boxs and is protected from light culture 2h.The present invention is handled using lysate, destroys the cell membrane of pollen so that coloring agent can fast and effeciently enter cell.By the present invention in that with lysate and coloring agent (Fluo 4AM ester), 20min is handled to pollen tube under room temperature (25 DEG C), calcium ion in pollen tube can successfully be dyed.
Description
Technical field
The present invention relates to a kind of methods that the polar growth field of pollen tube more particularly to pollen tube load fluorescent dye.
Background technology
The normal growth of pollen tube is multi-signal molecular regulation as a result, and calcium ion plays important work wherein
With.The higher calcium gradient in pollen tube cytoplasm tip is had observed that in various plants, the disappearance of calcium concentration gradient can cause to spend
Tube cell growth stops.Pollen tube top high calcium gradient may have following functions：First, it may participate in the adjusting of vesica secretion,
Because the calcium of high concentration is conducive to merging for Secretory vesicles and plasma membrane, outer row is promoted to act on, extends pollen tube；Second, high concentration
Calcium makes depolymerization of microfilaments, and the movement of organelle slows down, so calcium gradient can influence organelle by controlling the assembling of microfilament
Transport；Third, the calcium of high concentration is related with pollen tube oriented growth, and the free calcium ions of pollen tube top high concentration can inhibit flesh
The movement of filamentous actin-myosin regulation and control precipitates vesica, if increasing cytosolic free calcium concentration in pollen tube side, can make flower
Tube cell changes the original direction of growth, is grown to high calcium concentration side.For the effect of calcium ion in further Study of Pollens pipe,
We can use fluorescent labelling techniques.Fluorescence indicator is indispensable mark substance in this course, in plant flowers
Increasingly important role is played in tube cell construction and growth characteristics research.Currently used fluorescence indicator, which has, not to be destroyed
Eucaryotic cell structure under the premise of maintaining tissue activity, passes through fluorescence microscope, laser confocal microscope and flow cytometer etc.
The measure of feature instrument can obtain the specific fine structure of pollen tube and histofluorescence image.The success of this method will be based on
Dyestuff is successfully loaded into pollen (pipe), at present, the type of calcium ion fluorescent has：Single wavelength fluorescence probe, including
Fluo-3, quin-2, Calcium Green, Calcium Orange etc.；With double wave length fluorescent probe, mainly have indo-1,
Fura-2, fura-red etc..The stowage of calcium ion fluorescence indicator has：Electroporation (this method is big to cellular damage, if
Standby expensive, do not have to typically now), (this method has cell a degree of damage, in addition complicated for operation, work for microinjection
Amount is big, success rate is low, be not suitable for the smaller cell of individual, is not suitable for the shortcomings of cell batch all limits this method
Extensive use, patch-clamp (speed is slow, equipment is expensive), low temperature process (time is long), high temperature method (influencing survival).More than overcoming
Deficiency, need explore fast and effeciently load fluorescent dye method.
Invention content
In order to overcome the defects of the prior art, a kind of method that pollen tube loads fluorescent dye is provided.
The present invention is realized by following proposal：
A kind of method that pollen tube loads fluorescent dye, includes the following steps：
Pollen is wrapped in pan paper by the first step, is positioned in closed silica gel bottle and is stored in low temperature refrigerator；
Second step after pollen takes out from refrigerator, is added in after placing 2h at normal temperatures in culture solution, after being stirred for uniformly
It is placed in 25 DEG C of insulating boxs and cultivates 2h,
Third walks, and by culture solution and lysate, coloring agent is corresponding mixes, and is protected from light 10-30min, is subsequently placed in 25 DEG C of constant temperature
Culture 2h is protected from light in case.
In the second step, the culture solution is 2- (N- morpholines) ethanesulfonic acid monohydrate that solute is dissolved in 30mM
In solution, the solute includes sucrose, polyethylene glycol, boric acid calcium nitrate；
Wherein, the mass percent of solute is as follows in the culture solution：10% sucrose, 20% polyethylene glycol,
0.01% boric acid, 0.03% calcium nitrate.
The volume ratio of the culture solution and lysate, coloring agent is 48:1:1.
A concentration of 100mM that CTAB mass percents in the lysate are 2%, Tris-HCl, EDTA's is a concentration of
40mM。
The coloring agent is acetoxymethyl ester derivative, can with penetration cell film, into cell after can be by intracellular esterase institute water
Solution, the Fluo-4 of generation then can with calcium binding and send out fluorescence.
Beneficial effects of the present invention are：
The coloring agent (Fluo-4 AM-ester) that the method that a kind of pollen tube of the present invention loads fluorescent dye uses is a kind of
It can be with the fluorescent dye of penetration cell film, mainly by the calcium binding that dissociates with intracellular and iuntercellular to Ca2+Signal point
Cloth carries out quantitative analysis, it sensitive can reflect the variation of free intracellular calcium concentration, and coloring agent enters after cell through non-spy
Different in nature esterase is sloughed AM and is stayed in into the cell as Fluo-4, its fluorescence intensity is itself after Fluo-4 is combined with intracellular free calcium
More than 40 times or more.When studying intracellular calcium using calcium ion fluorescent combination fluorescence analysis, fluorescent dye
Loading be committed step in this method, the present invention is handled using lysate, destroys the cell membrane of pollen so that coloring agent
Cell can fast and effeciently be entered.By the present invention in that with lysate and coloring agent (Fluo-4AM-ester), in room temperature (25
DEG C) under to pollen tube handle 20min, calcium ion in pollen tube can successfully be dyed.
Specific embodiment
With reference to specific embodiment, the present invention is further described：
In the present invention, mM is substance withdrawl syndrome unit, is bold and unconstrained mole every liter.
A kind of method that pollen tube loads fluorescent dye, includes the following steps：
Pollen is wrapped in pan paper by the first step, is positioned in closed silica gel bottle and is stored in low temperature refrigerator；
Second step after pollen takes out from refrigerator, is added in after placing 2h at normal temperatures in culture solution, after being stirred for uniformly
It is placed in 25 DEG C of insulating boxs and cultivates 2h,
Third walks, and by culture solution and lysate, coloring agent is corresponding mixes, and is protected from light 10-30min, is subsequently placed in 25 DEG C of constant temperature
Culture 2h is protected from light in case.
In the second step, the culture solution is 2- (N- morpholines) ethanesulfonic acid monohydrate that solute is dissolved in 30mM
In solution, the solute includes sucrose, polyethylene glycol, boric acid calcium nitrate；
Wherein, the mass percent of solute is as follows in the culture solution：10% sucrose, 20% polyethylene glycol,
0.01% boric acid, 0.03% calcium nitrate.
The volume ratio of the culture solution and lysate, coloring agent is 48:1:1.
A concentration of 100mM that CTAB mass percents in the lysate are 2%, Tris-HCl, EDTA's is a concentration of
40mM。
The coloring agent is acetoxymethyl ester derivative, can with penetration cell film, into cell after can be by intracellular esterase institute water
Solution, the Fluo-4 of generation then can with calcium binding and send out fluorescence.
Wherein the molecular formula of Fluo-4 is C51H50F2N2O23, molecular weight 1096.94.
In actual use, also LaCl can be added in pollen solution after the completion of third step3, can make to dissociate in pollen tube
Calcium ion concentration dramatically increases.
The coloring agent (Fluo-4 M-ester) that the method that a kind of pollen tube of the present invention loads fluorescent dye uses is a kind of
It can be with the fluorescent dye of penetration cell film, mainly by the calcium binding that dissociates with intracellular and iuntercellular to Ca2+Signal point
Cloth carries out quantitative analysis, it sensitive can reflect the variation of free intracellular calcium concentration, and coloring agent enters after cell through non-spy
Different in nature esterase is sloughed AM and is stayed in into the cell as Fluo-4, its fluorescence intensity is itself after Fluo-4 is combined with intracellular free calcium
More than 40 times or more.When studying intracellular calcium using calcium ion fluorescent combination fluorescence analysis, fluorescent dye
Loading be committed step in this method, the present invention is handled using lysate, destroys the cell membrane of pollen so that coloring agent
Cell can fast and effeciently be entered.By the present invention in that with lysate and coloring agent (Fluo-4AM-ester), in room temperature (25
DEG C) under to pollen tube handle 20min, calcium ion in pollen tube can successfully be dyed.
Although more detailed elaboration is done to technical scheme of the present invention and has been enumerated, it should be understood that for ability
For field technique personnel, modification is made to above-described embodiment or uses equivalent alternative solution, this is to those skilled in the art
It is it is clear that these modifications or improvements without departing from theon the basis of the spirit of the present invention, belong to the present invention for member
Claimed range.
Claims (2)
1. a kind of method that pollen tube loads fluorescent dye, which is characterized in that include the following steps：
Pollen is wrapped in pan paper by the first step, is positioned in closed silica gel bottle and is stored in low temperature refrigerator；
Second step after pollen takes out from refrigerator, adds in culture solution after placing 2h at normal temperatures, is stirred for uniformly being placed on
2 h are cultivated in 25 DEG C of insulating boxs,
Third walks, and by culture solution and lysate, coloring agent is corresponding mixes, and is protected from light 10-30min, is subsequently placed in 25 DEG C of insulating boxs
It is protected from light 2 h of culture；
In the second step, the culture solution be solute is dissolved in 30 mM 2- (N- morpholines) ethanesulfonic acid monohydrate it is molten
In liquid, the solute includes sucrose, polyethylene glycol, boric acid, calcium nitrate；
Wherein, the mass percent of solute is as follows in the culture solution：10% sucrose, 20% polyethylene glycol, 0.01%
Boric acid, 0.03% calcium nitrate；
A concentration of 100 mM that CTAB mass percents in the lysate are 2%, Tris-HCl, a concentration of the 40 of EDTA
mM；
The coloring agent is Fluo-4 AM-ester.
2. the method that a kind of pollen tube according to claim 1 loads fluorescent dye, it is characterised in that：The culture solution with
Lysate, coloring agent volume ratio be 48:1:1.
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| CN201510944828.7A CN105486565B (en) | 2015-12-16 | 2015-12-16 | A kind of method that pollen tube loads fluorescent dye |
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|---|---|---|---|
| CN201510944828.7A CN105486565B (en) | 2015-12-16 | 2015-12-16 | A kind of method that pollen tube loads fluorescent dye |
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| CN105486565A CN105486565A (en) | 2016-04-13 |
| CN105486565B true CN105486565B (en) | 2018-07-03 |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108680418B (en) * | 2018-06-01 | 2021-03-23 | 广东金作农业科技有限公司 | Dyeing liquid and method for quickly dyeing cruciferous crop pollen |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101398381A (en) * | 2008-11-12 | 2009-04-01 | 南京农业大学 | Fluorescence labeling method for pear pollen and pollen tube fibril framework |
| CN101477000A (en) * | 2009-01-05 | 2009-07-08 | 南京农业大学 | Fluorescence labeling method for cotton pollen tube microfilament framework |
| CN104531824A (en) * | 2014-12-16 | 2015-04-22 | 北京农学院 | Labeling method of apple pollen tube microfilament |
-
2015
- 2015-12-16 CN CN201510944828.7A patent/CN105486565B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101398381A (en) * | 2008-11-12 | 2009-04-01 | 南京农业大学 | Fluorescence labeling method for pear pollen and pollen tube fibril framework |
| CN101477000A (en) * | 2009-01-05 | 2009-07-08 | 南京农业大学 | Fluorescence labeling method for cotton pollen tube microfilament framework |
| CN104531824A (en) * | 2014-12-16 | 2015-04-22 | 北京农学院 | Labeling method of apple pollen tube microfilament |
Non-Patent Citations (4)
| Title |
|---|
| Disarrangement of actin filaments and Ca2+ gradient by CdCl2 alters cell wall construction in Arabidopsis thaliana root hairs by inhibiting vesicular trafficking;Jun-Ling Fan et al.;《Journal of Plant Physiology》;20111231;第168卷;第1157-1167页 * |
| 利用荧光探针Fluo-4检测胸腺细胞内钙离子浓度变化;于力方 等;《标记免疫分析与临床》;20050930;第12卷(第3期);第161-163页 * |
| 百合花粉细胞中钙离子的荧光测定方法;尚忠林 等;《植物生理学通讯》;20010831;第37卷(第4期);第319-322页 * |
| 青杆花粉管生长过程中囊泡运输与Ca2+分布的细胞学研究;李伶俐 等;《电子显微学报》;20100430;第29卷(第2期);第163-164页 * |
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| CN105486565A (en) | 2016-04-13 |
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