CN101845415B - Method for separating Casparian strip - Google Patents
Method for separating Casparian strip Download PDFInfo
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- CN101845415B CN101845415B CN2010101786013A CN201010178601A CN101845415B CN 101845415 B CN101845415 B CN 101845415B CN 2010101786013 A CN2010101786013 A CN 2010101786013A CN 201010178601 A CN201010178601 A CN 201010178601A CN 101845415 B CN101845415 B CN 101845415B
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- 238000000034 method Methods 0.000 title claims abstract description 28
- 239000007788 liquid Substances 0.000 claims abstract description 37
- 108090000790 Enzymes Proteins 0.000 claims abstract description 24
- 102000004190 Enzymes Human genes 0.000 claims abstract description 24
- 229940088598 enzyme Drugs 0.000 claims abstract description 24
- 108010059892 Cellulase Proteins 0.000 claims abstract description 14
- 229940106157 cellulase Drugs 0.000 claims abstract description 14
- 108010059820 Polygalacturonase Proteins 0.000 claims abstract description 13
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 claims abstract description 8
- KRVSOGSZCMJSLX-UHFFFAOYSA-L chromic acid Substances O[Cr](O)(=O)=O KRVSOGSZCMJSLX-UHFFFAOYSA-L 0.000 claims abstract description 8
- 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 8
- 229910017604 nitric acid Inorganic materials 0.000 claims abstract description 8
- 241000196324 Embryophyta Species 0.000 claims description 44
- 235000007164 Oryza sativa Nutrition 0.000 claims description 39
- 235000009566 rice Nutrition 0.000 claims description 38
- 238000005204 segregation Methods 0.000 claims description 16
- 239000007979 citrate buffer Substances 0.000 claims description 14
- 239000007864 aqueous solution Substances 0.000 claims description 7
- 239000011259 mixed solution Substances 0.000 claims description 7
- 238000007654 immersion Methods 0.000 claims description 6
- 240000007594 Oryza sativa Species 0.000 claims description 2
- 238000002791 soaking Methods 0.000 claims description 2
- 238000000926 separation method Methods 0.000 abstract description 8
- 239000000203 mixture Substances 0.000 abstract description 6
- 230000000694 effects Effects 0.000 abstract description 2
- 108010093305 exopolygalacturonase Proteins 0.000 abstract 1
- 241000209094 Oryza Species 0.000 description 37
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 21
- 239000008367 deionised water Substances 0.000 description 18
- 229910021641 deionized water Inorganic materials 0.000 description 15
- 238000011161 development Methods 0.000 description 9
- 230000018109 developmental process Effects 0.000 description 9
- 239000000463 material Substances 0.000 description 7
- 239000000126 substance Substances 0.000 description 5
- 238000012137 double-staining Methods 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- 238000000862 absorption spectrum Methods 0.000 description 3
- 210000004027 cell Anatomy 0.000 description 3
- 238000001035 drying Methods 0.000 description 3
- 230000005284 excitation Effects 0.000 description 3
- 235000015097 nutrients Nutrition 0.000 description 3
- 230000007226 seed germination Effects 0.000 description 3
- 244000025254 Cannabis sativa Species 0.000 description 2
- 108010029182 Pectin lyase Proteins 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 230000005540 biological transmission Effects 0.000 description 2
- 229920002678 cellulose Polymers 0.000 description 2
- 239000001913 cellulose Substances 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000004615 ingredient Substances 0.000 description 2
- 238000011835 investigation Methods 0.000 description 2
- IOLCXVTUBQKXJR-UHFFFAOYSA-M potassium bromide Chemical compound [K+].[Br-] IOLCXVTUBQKXJR-UHFFFAOYSA-M 0.000 description 2
- 241000209510 Liliopsida Species 0.000 description 1
- 229930183415 Suberin Natural products 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 210000003725 endotheliocyte Anatomy 0.000 description 1
- 210000002615 epidermis Anatomy 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- HZVOZRGWRWCICA-UHFFFAOYSA-N methanediyl Chemical compound [CH2] HZVOZRGWRWCICA-UHFFFAOYSA-N 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 230000035515 penetration Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000010129 solution processing Methods 0.000 description 1
- 230000008719 thickening Effects 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 230000002792 vascular Effects 0.000 description 1
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/005—Methods for micropropagation; Vegetative plant propagation using cell or tissue culture techniques
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- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/04—Plant cells or tissues
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Abstract
The invention discloses a method for separating a Casparian strip, which comprises the following steps that: 1) a plant primary root is soaked with a segregating liquid, and the soaked plant primary root is recorded as a segregated plant primary root; 2) the segregated plant primary root is enzymolyzed, and the segregated plant primary root is recorded as an enzymolyzed plant primary root; and 3) the enzymolyzed plant primary root is separated to obtain the Casparian strip, wherein the segregating liquid is of an isometric liquid mixture of (7-12)% ( mass percentage) of aqueous chromic acid solution and (7-12)% (mass percentage) of aqueous nitric acid solution. The invention has the following advantages: 1) the operation is simple and fast, and the separation time is shortened; 2) cellulase and pectinase are used for moderate enzymolysis, so the use amount of enzyme is reduced, and the cost is saved; and 3) the separation effect is good.
Description
Technical field
The present invention relates to a kind of method of separating Casparian strip in the biological technical field.
Background technology
Casparian strip is the suberification and the lignified banded thickening of higher plant tegumental cell radial wall and transverse wall, and major function is to stop moisture to tissue penetration, is controlling the matter transportation between cortex and the vascular cylinder.Its width has bigger difference with different plants.Whether Casparian strip mainly is distributed in the endodermis and the exodermis of primary root, in the living organ of gas such as stem, leaf, exist then still disputable.Casparian strip mainly influences the ectoplast transport pathway in the transportation of plant root water and inorganic nutrients, that is water and inorganic nutrients are via matter transmission transportation between the cell walls of epidermis, exodermis and tegumental cell.When water and inorganic nutrients are come the endodermis cell, change by matter transmission between the cytolemma of endotheliocyte, finally get into intrastelar tracheid because of the waterproof character of Casparian strip.
For a long time, the definite chemical ingredients of Casparian strip never obtains further investigation, and major cause is also not find a kind of simple possible, and can obtain the experimental technique of a large amount of Casparian strips.Usual earlier uses natural enzymolysis to separate endodermis, but its efficient very low (several milligram), and enzymolysis time long (more than 10 days).
Summary of the invention
The purpose of this invention is to provide a kind of method of separating Casparian strip.
The method of separation Casparian strip provided by the invention may further comprise the steps: 1) soak the plant primary root with segregation liquid, the plant primary root note after soaking is made segregation back plant primary root; 2) said segregation back plant primary root is carried out enzymolysis, the note of the plant primary root behind the enzymolysis is made plant primary root behind the enzymolysis; 3) behind said enzymolysis, separate the plant primary root and obtain Casparian strip; Said segregation liquid is (7-12) % (mass percent) chromic acid aqueous solution and (7-12) the equal-volume mixed solution of % (mass percent) aqueous nitric acid; The temperature of said immersion is (24-26) ℃, and the time of said immersion is (20-30) minute.
The enzyme liquid that said enzymolysis adopts is for obtaining enzyme liquid with (12-32) U polygalacturonase with (18-48) in the U cellulase adding 2ml citrate buffer; The concentration of said citrate buffer is (0.01-0.05) mol/L, and the pH value is 3.0; The temperature of said enzymolysis is (24-26) ℃, and the time of said enzymolysis is (3-4) day.
Said segregation liquid is preferably the equal-volume mixed solution of 7%, 10% or 12% (mass percent) chromic acid aqueous solution and 7%, 10% or 12% (mass percent) aqueous nitric acid; The temperature of said immersion is preferably 24 ℃, 25 ℃ or 26 ℃, and said soak time is preferably 20 minutes, 25 minutes or 30 minutes.
Said enzyme liquid is with obtaining enzyme liquid in 12U, 20U or 32U polygalacturonase and 18U, 30U or the 48U cellulase adding 2ml citrate buffer; The concentration of said citrate buffer is preferably 0.01mol/L, 0.03mol/L or 0.05mol/L; The temperature of said enzymolysis is preferably 24 ℃, 25 ℃ or 26 ℃, and the time is preferably 3 days, 3.5 days or 4 days.
Said cellulase is specially cellulase R-10, and English name is: cellulose " Onozuka " R-10, and glad through Bioisystech Co., Ltd of section available from Beijing, PIN is 6008; Said polygalacturonase is specially polygalacturonase Y-23, and English name is Pectolyase Y-23, and glad available from Beijing is 6038 through Bioisystech Co., Ltd of section PIN.
Said enzymolysis process is generally whenever changed once said enzyme liquid at a distance from 1 day.
Said method also is included in the preceding step that plant primary root behind the said enzymolysis is vibrated of step 3); The time of said vibration is generally (4-5) hour; Be preferably 4 hours, 4.5 hours or 5 hours; Shaking speed is generally (50-60) rpm, is preferably 50rpm, 55rpm or 60rpm, and rotation radius is 50mm.
Said plant primary root is that length is the nascent root segment of plant of (0.5-1) cm, is preferably the nascent root segment of plant of 0.5cm, 0.8cm or 1cm.
Said plant can be monocotyledons, is preferably grass, especially is preferably paddy rice.
Said paddy rice can be the paddy rice of following kind: the Liao Dynasty's drought 109, the fine or field rich 202 of Japan.
Casparian strip separation method provided by the invention, it has the following advantages: 1) simple to operate, quick: as to have shortened separation time; 2) re-use cellulase after acid solution is handled and polygalacturonase carries out gentle enzymolysis, reduced the usage quantity of enzyme, provide cost savings; 3) good separating effect: can under fluorescent microscope, clearly observe root endodermis Casparian strip through the blue double staining of Berberine-aniline.
Experiment showed, the extraction time weak point that adopts method provided by the invention to obtain Casparian strip, can accomplish about 3-5 days greatly; The consumption of cellulase reduces, and reaches cost-effective purpose; For fine structure and the chemical ingredients of furtheing investigate Casparian strip provides enough materials.This shows that the method that the acid solution processing combines the enzymolysis technical point to leave Casparian strip is that the further investigation of higher plant Casparian strip is laid a good foundation, and especially in the research of unifacial leaf grass root endodermis Casparian strip, plays a significant role, actual application value is high.
Description of drawings
Fig. 1 is the structure of the non-irrigated 109 rice root endodermis Casparian strips of observed the Liao Dynasty under fluorescent microscope
Fig. 2 is the structure of the fine rice root endodermis of observed Japan Casparian strip under fluorescent microscope
Fig. 3 is the structure of the rich 202 rice root endodermis Casparian strips in observed field under fluorescent microscope
Fig. 4 is 3 kinds of paddy rice (the Liao Dynasty's drought 109, Japanese warm and fine field rich 202) root endodermis Casparian strip FFIRs
Embodiment
Used material, reagent etc. like no specified otherwise, all can obtain from commercial sources among the following embodiment.
Method therefor is ordinary method if no special instructions among the following embodiment, and said percentage composition is the quality percentage composition if no special instructions.
The separation of embodiment 1, rice root endodermis Casparian strip and microscopic and chemical composition analysis
Paddy rice (the Liao Dynasty's drought 109) seed (Liaoning Panjin north agrotechnique development corporation, Ltd.) in 20-25 ℃ of following vernalization, behind the seed germination, is put in 25 ± 2 ℃ of greenhouses and carried out the water planting plantation with 1/2 * Hoagland substratum, be cultured to tri-leaf period, get the primary root of seedling.
1) above-mentioned primary root is cut into the long segment of 0.5cm, with deionized water wash to the substratum of removing on the primary root;
2) 20 sections primary roots that step 1) obtained are put into 2ml segregation liquid, after being pumped to material and sinking, soak 25 minutes to the transparent beige that is of the tip of a root at 24 ℃, with deionized water rinsing 2-3 time, and plant primary root after obtaining to emanate.Said segregation liquid is the equal-volume mixed solution of 10% (mass percent) chromic acid aqueous solution and 10% (mass percent) aqueous nitric acid;
3) will emanate back plant primary root put into the mid-24 ℃ of following enzymolysis of enzyme liquid after 3 days with plant primary root behind 2-3 acquisition of the deionized water rinsing enzymolysis; Whenever changed enzyme liquid once at a distance from 1 day in the enzymolysis process; Said enzyme liquid is with obtaining enzyme liquid in 20U polygalacturonase and the 30U cellulase adding 2ml citrate buffer; The concentration of said citrate buffer is 0.01mol/L.
Said cellulase is specially cellulase R-10, and English name is: cellulose " Onozuka " R-10, and glad through Bioisystech Co., Ltd of section available from Beijing, PIN is 6008; Said polygalacturonase is specially polygalacturonase Y-23, and English name is Pectolyase Y-23, and glad available from Beijing is 6038 through Bioisystech Co., Ltd of section PIN.
4) place deionized water on shaking table, to vibrate 4 hours plant primary root behind the enzymolysis, shake fast 50rpm, rotation radius is 50mm;
5) under anatomical lens, from plant primary root behind the enzymolysis of step 4), separate and obtain paddy rice (the Liao Dynasty's drought 109) root endodermis Casparian strip;
6) with paddy rice (the Liao Dynasty drought 109) root endodermis Casparian strip with washed with de-ionized water 2-3 time, be kept in the deionized water, frozen drying, subsequent use.
Adopting uses the same method obtains root endodermis Casparian strip from paddy rice (Japan is fine) (Liaoning Panjin north agrotechnique development corporation, Ltd.) and paddy rice (field rich 202) (Liaoning Panjin north agrotechnique development corporation, Ltd.) respectively.
The root endodermis Kai Shi of the paddy rice of above-mentioned acquisition (the Liao Dynasty's drought 109), paddy rice (Japan is fine) and paddy rice (field rich 202) is used the blue double staining of Berberine-aniline respectively; Be to observe under the fluorescent microscope of 365nm at the ultraviolet excitation wavelength; The result is respectively shown in Fig. 1-3; Can find out that from Fig. 1-3 Casparian strip has reticulated structure, prove to separate to obtain Casparian strip.
The separation of embodiment 2, rice root endodermis Casparian strip and microscopic and chemical composition analysis
Paddy rice (the Liao Dynasty's drought 109) seed (Liaoning Panjin north agrotechnique development corporation, Ltd.) in 20-25 ℃ of following vernalization, behind the seed germination, is put in 25 ± 2 ℃ of greenhouses and carried out the water planting plantation with 1/2 * Hoagland substratum, be cultured to tri-leaf period, get the primary root of seedling.
1) above-mentioned primary root is cut into the long segment of 0.8cm, with deionized water wash to the substratum of removing on the primary root;
2) 20 sections primary roots that step 1) obtained are put into 2ml segregation liquid, after being pumped to material and sinking, soak 20 minutes to the transparent beige that is of the tip of a root at 25 ℃, with deionized water rinsing 2-3 time, and plant primary root after obtaining to emanate.Said segregation liquid is the equal-volume mixed solution of 7% (mass percent) chromic acid aqueous solution and 7% (mass percent) aqueous nitric acid;
3) will emanate back plant primary root put into the rearmounted 25 ℃ of following enzymolysis of enzyme liquid after 3.5 days with plant primary root behind 2-3 acquisition of the deionized water rinsing enzymolysis; Whenever changed enzyme liquid once at a distance from 1 day in the enzymolysis process; Said enzyme liquid is with obtaining enzyme liquid in 12U polygalacturonase and the 18U cellulase adding 2ml citrate buffer; The concentration of said citrate buffer is 0.03mol/L.
4) place deionized water on shaking table, to vibrate 4.5 hours plant primary root behind the enzymolysis then, shake fast 55rpm, rotation radius is 50mm;
5) under anatomical lens, from plant primary root behind the enzymolysis of step 4), separate and obtain paddy rice (the Liao Dynasty's drought 109) root endodermis Casparian strip;
6) with paddy rice (the Liao Dynasty drought 109) root endodermis Casparian strip with washed with de-ionized water 2-3 time, be kept in the deionized water, frozen drying, subsequent use.
Adopting uses the same method obtains root endodermis Casparian strip from paddy rice (Japan is fine) (Liaoning Panjin north agrotechnique development corporation, Ltd.) and paddy rice (field rich 202) (Liaoning Panjin north agrotechnique development corporation, Ltd.) respectively.
The root endodermis Kai Shi of the paddy rice of above-mentioned acquisition (the Liao Dynasty's drought 109), paddy rice (Japan is fine) and paddy rice (field rich 202) is used the blue double staining of Berberine-aniline respectively; Be to observe under the fluorescent microscope of 365nm at the ultraviolet excitation wavelength; The result can find out all that with consistent shown in Fig. 1-3 Casparian strip has reticulated structure.
The separation of embodiment 3, rice root endodermis Casparian strip and microscopic and chemical composition analysis
Paddy rice (the Liao Dynasty's drought 109) seed (Liaoning Panjin north agrotechnique development corporation, Ltd.) in 20-25 ℃ of following vernalization, behind the seed germination, is put in 25 ± 2 ℃ of greenhouses and carried out the water planting plantation with 1/2 * Hoagland substratum, be cultured to tri-leaf period, get the primary root of seedling.
1) above-mentioned primary root is cut into the long segment of 1cm, with deionized water wash to the substratum of removing on the primary root;
2) 20 sections primary roots that step 1) obtained are put into 2ml segregation liquid, after being pumped to material and sinking, soak 30 minutes to the transparent beige that is of the tip of a root at 26 ℃, with deionized water rinsing 2-3 time, and plant primary root after obtaining to emanate.Said segregation liquid is the equal-volume mixed solution of 12% (mass percent) chromic acid aqueous solution and 12% (mass percent) aqueous nitric acid;
3) will emanate back plant primary root put into the rearmounted 26 ℃ of following enzymolysis of enzyme liquid after 4 days with plant primary root behind 2-3 acquisition of the deionized water rinsing enzymolysis; Whenever changed enzyme liquid once at a distance from 1 day in the enzymolysis process; Said enzyme liquid is with obtaining enzyme liquid in 32U polygalacturonase and the 48U cellulase adding 2ml citrate buffer; The concentration of said citrate buffer is 0.05mol/L.
4) place deionized water on shaking table, to vibrate 5 hours plant primary root behind the enzymolysis then, shake fast 60rpm, rotation radius is 50mm;
5) under anatomical lens, from plant primary root behind the enzymolysis of step 4), separate and obtain paddy rice (the Liao Dynasty's drought 109) root endodermis Casparian strip;
6) with paddy rice (the Liao Dynasty drought 109) root endodermis Casparian strip with washed with de-ionized water 2-3 time, be kept in the deionized water, frozen drying, subsequent use.
Adopt and use the same method respectively from paddy rice (Japan is fine) (Liaoning Panjin north agrotechnique development corporation, Ltd.; Business Name please offer for sale) and paddy rice (field rich 202) (Liaoning Panjin north agrotechnique development corporation, Ltd., Business Name please offer for sale) acquisition root endodermis Casparian strip.
The root endodermis Kai Shi of the paddy rice of above-mentioned acquisition (the Liao Dynasty's drought 109), paddy rice (Japan is fine) and paddy rice (field rich 202) is used the blue double staining of Berberine-aniline respectively; Be to observe under the fluorescent microscope of 365nm at the ultraviolet excitation wavelength; The result is consistent with Fig. 1-3, can find out that all Casparian strip has reticulated structure.
The infrared absorption spectrum of embodiment 4, rice root endodermis Casparian strip
The root endodermis Casparian strip of three kind paddy rice that will be obtained by embodiment 1 is used pressing potassium bromide troche; Under Fourier transformation infrared spectrometer, measure infrared absorption spectrum respectively, the result is as shown in Figure 4, and Liaohan109 is the Liao Dynasty's drought 109; Oryzasativa cv.Nipponbare is that Japan is fine; Tianfeng202 is that the field is rich 202, because suberin is the important component of Casparian strip, and methylene radical CH
2The absorption peak of unsymmetrically and symmetrical stretching vibration is corky charateristic avsorption band.And this charateristic avsorption band appears at 2855cm respectively in rich 202 outside organization of the Liao Dynasty drought 109, Japanese warm and fine field
-1, 2854cm
-And 2854cm
-1As reference, in the Liao Dynasty's drought 109, the rich 202 internal layer Casparian strips in Japanese warm and fine field, appear at 2854cm respectively
-1, 2854cm
-1And 2855cm
-1The peak, therefore can prove that extracting the material that obtains is Casparian strip.
Adopting uses the same method detects the root endodermis Casparian strip with three kind paddy rice of embodiment 3 acquisitions by embodiment 2, and infrared absorption spectrum is all consistent with Fig. 4, and therefore proof is extracted the material that obtains and is Casparian strip too.
Claims (8)
1. method of separating Casparian strip may further comprise the steps:
1) soaks the plant primary root with segregation liquid, the plant primary root note after soaking is made segregation back plant primary root;
2) said segregation back plant primary root is carried out enzymolysis, the note of the plant primary root behind the enzymolysis is made plant primary root behind the enzymolysis;
3) behind said enzymolysis, separate the plant primary root and obtain Casparian strip;
Said segregation liquid is mixed solution that to be the chromic acid aqueous solution of 7-12% by mass percent obtain with aqueous nitric acid equal-volume mixing that mass percent is 7-12%; The temperature of said immersion is 24-26 ℃, and the time of said immersion is 20-30 minute;
The enzyme liquid that said enzymolysis adopts is with obtaining enzyme liquid in 12-32U polygalacturonase and the 18-48U cellulase adding 2ml citrate buffer; The concentration of said citrate buffer is 0.01-0.05mol/L, and the pH value is 3.0; The temperature of said enzymolysis is 24-26 ℃, and the time of said enzymolysis is 3-4 days;
Said plant is a paddy rice.
2. method according to claim 1 is characterized in that: said segregation liquid is that mass percent is that 7%, 10% or 12% chromic acid aqueous solution and mass percent are the equal-volume mixed solution of 7%, 10% or 12% aqueous nitric acid; The temperature of said immersion is 24 ℃, 25 ℃ or 26 ℃, and said soak time is 20 minutes, 25 minutes or 30 minutes.
3. method according to claim 1 and 2 is characterized in that: said enzyme liquid is with obtaining enzyme liquid in 12U, 20U or 32U polygalacturonase and 18U, 30U or the 48U cellulase adding 2ml citrate buffer; The concentration of said citrate buffer is 0.01mol/L, 0.03mol/L or 0.05mol/L; The temperature of said enzymolysis is 24 ℃, 25 ℃ or 26 ℃, and said enzymolysis time is 3 days, 3.5 days or 4 days.
4. method according to claim 1 is characterized in that: said enzymolysis was for whenever changing once said enzyme liquid at a distance from 1 day.
5. method according to claim 1 is characterized in that: said method also is included in the preceding step that plant primary root behind the said enzymolysis is vibrated of step 3), and the time of said vibration is 4-5 hour, shakes speed and is 50-60rpm, and rotation radius is 50mm.
6. method according to claim 5 is characterized in that: the time of said vibration is 4 hours, 4.5 hours or 5 hours, shakes speed and is 50rpm, 55rpm or 60rpm.
7. method according to claim 1 is characterized in that: said plant primary root is that length is the nascent root segment of plant of 0.5-1cm.
8. method according to claim 7 is characterized in that: the plant that said plant primary root is 0.5cm, 0.8cm or the 1cm root segment of coming into being.
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010101786013A CN101845415B (en) | 2010-05-17 | 2010-05-17 | Method for separating Casparian strip |
| PCT/CN2011/000431 WO2011143922A1 (en) | 2010-05-17 | 2011-03-16 | Process for the separation of casparian strip |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010101786013A CN101845415B (en) | 2010-05-17 | 2010-05-17 | Method for separating Casparian strip |
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| CN101845415A CN101845415A (en) | 2010-09-29 |
| CN101845415B true CN101845415B (en) | 2012-06-06 |
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| CN101845415B (en) * | 2010-05-17 | 2012-06-06 | 中国科学院植物研究所 | Method for separating Casparian strip |
| CN106119184B (en) * | 2016-01-19 | 2019-08-09 | 北京林业大学 | A method of separation casparian strip |
| CN116380595A (en) * | 2023-05-04 | 2023-07-04 | 生态环境部南京环境科学研究所 | Simple and effective bamboo leaf epidermis flaking method |
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Non-Patent Citations (4)
| Title |
|---|
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