CN101131385A - Rotifer Brachionus plicatilis microkernel preparation technology - Google Patents
Rotifer Brachionus plicatilis microkernel preparation technology Download PDFInfo
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- CN101131385A CN101131385A CNA2007101324493A CN200710132449A CN101131385A CN 101131385 A CN101131385 A CN 101131385A CN A2007101324493 A CNA2007101324493 A CN A2007101324493A CN 200710132449 A CN200710132449 A CN 200710132449A CN 101131385 A CN101131385 A CN 101131385A
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- wheel animalcule
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- brachionus plicatilis
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A20/00—Water conservation; Efficient water supply; Efficient water use
- Y02A20/20—Controlling water pollution; Waste water treatment
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Abstract
The invention is a preparation technology of Brachionus plicatilis microkernel. The water pollution (such as heavy metal cadmium) is dangerous to the Brachionus plicatilis. In cell, the cell nuclear fission of Brachionus plicatilis is influenced and the microkernel can be produced. The microkernel number can appraise the toxicity degree of pollution. The preparation method includes the rotifer toxicity, tabletting, coloration and observation. The figure in abstract shows the size and form of the Brachionus plicatilis microkernel.
Description
Technical field:
The present invention relates to the preparation method of wheel animalcule micronucleus and, be that a kind of application cell toxicology is carried out the technology that environmental contaminants toxicity detects as the application of pollutant toxicity index.
Background technology:
In chemical substance, have a lot of can cause chromosomal unusual, chemical substance is brought out chromosome abnormality and is shown as that chromosome diminution or part form and the micronucleus that occurs in the nucleus level, detects the probability that micronucleus occurs and can be used as a kind of index of weighing chemical substance toxicity.At present, both at home and abroad for concentrating on the following aspects from the biological reaction to environmental pollutants of cell toxicology angle research; The one, utilize plant root tip cell response situation to heavy metal toxicity in the mitosis process, contain heavy metal cadmium ion solution cultivation 24 hours as putting into root systems of plant such as the broad bean tip of a root, root top of onion, the garlic tips of a root, pair cell nuclear dyes then, observe and add up the probability of generation micronucleus, weigh the toxicity of heavy metal cadmium ion with this; The 2nd, utilize zooblast to do similar experiment, as utilize small white mouse marrow micronucleus in erythrocytes rate to come the toxicity of detection of contamination.So far do not see and utilize the planktonic nucleus of water body to be carried out the report that micronucleus detects as trying material.With regard to the water pollutant environment measuring, make full use of the planktonic organism of water body, especially animal plankton to detect the degree of pollution be a desirable index, at present from the cell toxicology angle, especially utilize the micronucleus technique of widespread use in the terrestrial organism to be used as detecting index and also be in space state.
Summary of the invention:
The present invention describes with water body planktonic organism brachionus plicatilis as a kind of micronucleus technique of weighing the harmful degree of water pollutant, the technology of preparing and the application in the water body environment monitoring thereof of wheel animalcule micronucleus have mainly been solved, main contents are for carrying out toxicity test to wheel animalcule earlier, utilize again in the pressed disc method trailing wheel worm's ovum mother cell and prepare micronucleus, as the index of weighing pollutant toxicity, is a kind of novel water body environment detection technique with the size of statistics micronuclear rates.
Description of drawings:
The small nut of lines indication is a micronucleus among the figure, and two big nuclears are two nucleus that form after the egg mother cell nuclear disintegration around it.
Embodiment:
The present invention can realize by following operation steps:
One, wheel animalcule is gathered and differentiates: contain water sampling the water sample of brachionus plicatilis from the field, consider through considering mesh screen, concentrate the liquid that contains wheel animalcule, draw single wheel animalcule with U trap, be positioned on the microslide, under anatomical lens (4 * 10=40 times of mirror), differentiate the kind of wheel animalcule, pick out brachionus plicatilis.
Two, wheel animalcule enlarged culture: from the water sample of above field acquisition, choose brachionus plicatilis about 1000 and put into 6 porocyte culture plates and carry out enlarged culture.Select the bait of limnetic chlorella as brachionus plicatilis in the incubation for use: when throwing something and feeding limnetic chlorella, the ratio of chlorella and wheel animalcule nutrient solution is 1: 50.
Three, the heavy metal cadmium ion is attacked the poison cultivation: above wheel animalcule is concentrated and cultivates after 2-3 days, it is divided into 8 groups puts into 12 well culture plates respectively and carries out cadmium ion and attack poison experiment, method is as follows: according to the measured concentration range of cadmium ion toxicity dose test, determine that eight experiment concentration of cadmium ions are 0.1mg/L, 0.2mg/L, 0.4mg/L, 0.8mg/L, 1.2mg/L, 2.4mg/L, 4.8mg/L and the experiment of original fluid control group.The cadmium ion treating fluid of getting above-mentioned concentration respectively is placed in 12 well culture plates for 2 milliliters, in each culture hole, put into about 200 wheel animalcules that vigor is stronger again, the limnetic chlorella that adds 0.5 milliliter then in each culture hole again, the pH value of regulating treating fluid at last reaches about 8.0 it.Culture plate is put into 25 ℃ of incubators, cultivated 24-48 hour.
Four, micronucleus microscopy and film-making are observed: draw each concentration cadmium ion wheel animalcule nutrient solution with suction pipe respectively and drop on the microslide, add the 10%Giemsa dyeing liquor and carried out visitain 15 minutes, add to be placed under 10 * 10 power microscopes behind the cover glass and push cover glass gently with dissecting needle, the variation of wheel animalcule form in the viewing microscope of one side at this moment, push cover glass on one side repeatedly, till having seen that liquid flows out from the wheel animalcule afterbody; Adjust field of microscope, observe the metamorphosis situation of all wheel animalcules under the cover glass, till utilizing dissecting needle that about wheel animalcule of about 80% is all extruded liquid from its afterbody.At this moment, wipe, blot cover glass liquid on every side gently, adjust microscope, utilize 10 * 100 times oily mirror to observe with filter paper.Can in the visual field, find that a large amount of oocyte nucleis that is in the nucleus splitting status occurs this moment, because wheel animalcule belongs to the planktonic organism of the lonely female life of main battalion, its reproductive capacity is very strong, so each wheel animalcule can observe the existence of the such egg mother cell about 15-30.Simultaneously, be at these egg mother cells around the nucleus of division stage, the existence of the dark state cellule nuclear of dyeing (being referred to as micronucleus) also can occur being in.
Five, micronucleus statistics and analysis: number and observed total oocyte number that micronucleus occurs are counted, observed 5000 cells under each concentration of cadmium ions approximately, statistics micronuclear rates, micronuclear rates (%)=micronucleus number/total inspection cell number.The micronuclear rates statistics sees Table 1
Table 1 wheel animalcule micronuclear rates statistics
| Concentration of cadmium ions mg/L | Average micronucleus number | Average micronucleus |
| 0 0.1 0.2 0.4 0.8 1.2 2.4 4.8 | 1 0.67 2.33 6 5 3 0.33 0.33 | 0.63 0.51 1.67 4.52 4.07 2.38 0.29 0.3 |
As can be seen from Table 1, cadmium ion with variable concentrations is handled wheel animalcule, the micronucleus probability that occurs in the wheel animalcule cell exists difference, and in the scope of concentration of cadmium ions from least concentration 0mg/L to 0.4mg/L, the micronuclear rates that is produced is and increases progressively trend, and in the scope of 0.4mg/L to 4.8mg/L, the wheel animalcule micronuclear rates trend that tapers off again.The former micronuclear rates incremental variations explanation gradually is very regular governed with the wheel animalcule micronuclear rates as the toxicity of weighing the heavy metal cadmium ion, and the probability that can this micronucleus occur is as the index of estimating water pollutant toxicity such as heavy metal; Latter's micronuclear rates is the variation of decline trend gradually again is because along with concentration of cadmium ions improves, its toxicity is strong more, has produced the toxicity inhibition, and the wheel animalcule oocyte nuclei no longer divides, and the normal fecundity of wheel animalcule has been subjected to disturbing and has caused.
Claims (1)
1. brachionus plicatilis micronucleus preparation method:
Utilize heavy metal such as cadmium-ion solution that brachionus plicatilis is carried out acute toxicity testing, handle and carry out the polypide bulk dyeing with the 10%Giemsa dyestuff after 24 hours, covered on wheel animalcule dyeed after 15 minutes; At 10 * 10=100 microscopically doubly, find wheel animalcule earlier, utilize dissecting needle to push cover glass downwards again in the position of wheel animalcule, strength is by gently increasing the weight of gradually, repeated multiple times until seeing that having nuclear egg mother cell is flowed out from both sides by wheel animalcule health afterbody, is squeezed to till the outflow fully; Utilize oily mirror, 10 * 100=1000 microscopically check oocyte nuclei is doubly found some little nucleus, i.e. micronucleus to occur near chief cell nuclear.
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| Application Number | Priority Date | Filing Date | Title |
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| CN2007101324493A CN101131385B (en) | 2007-09-17 | 2007-09-17 | Rotifer Brachionus plicatilis microkernel preparation technology |
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| CN2007101324493A CN101131385B (en) | 2007-09-17 | 2007-09-17 | Rotifer Brachionus plicatilis microkernel preparation technology |
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| CN101131385A true CN101131385A (en) | 2008-02-27 |
| CN101131385B CN101131385B (en) | 2010-11-10 |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102433371A (en) * | 2011-10-18 | 2012-05-02 | 南京师范大学 | Overall rotifer male fluorescent staining and in vitro sperm observation method |
| RU2461894C1 (en) * | 2011-07-12 | 2012-09-20 | Государственное образовательное учреждение высшего профессионального образования "Северо-Осетинская государственная медицинская академия" Министерства здравоохранения и социального развития РФ | Method for prevention of toxic action of cadmium in experimental animals suffering chronic poisoning |
| RU2462764C1 (en) * | 2011-07-13 | 2012-09-27 | Государственное образовательное учреждение высшего профессионального образования "Северо-Осетинская государственная медицинская академия" Министерства здравоохранения и социального развития РФ | Method for prevention of toxic action of cadmium in experimental animals suffering chronic poisoning |
| RU2462763C1 (en) * | 2011-07-13 | 2012-09-27 | Государственное образовательное учреждение высшего профессионального образования "Северо-Осетинская государственная медицинская академия" Министерства здравоохранения и социального развития РФ | Method for prevention of toxic action of cadmium in experimental animals suffering chronic poisoning |
| CN105076108A (en) * | 2015-07-07 | 2015-11-25 | 安徽师范大学 | Short-term fixing preservation method and long-term fixing preservation method for rotifer specimen |
| CN108485981A (en) * | 2018-03-27 | 2018-09-04 | 中国科学院城市环境研究所 | A kind of separation of arenaceous shell worm and culture technique |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1039117A (en) * | 1989-07-14 | 1990-01-24 | 复旦大学 | A kind of cell nucleus analysis and device that is used for the environmental pollution biological detection |
| DK1769081T3 (en) * | 2004-07-01 | 2016-03-14 | Litron Lab Ltd | PROCESS FOR COUNTING OF MAMMALS CELL micronuclei WITH A HEAD WEIGHT OF DIFFERENTIAL STAINING OF micronuclei AND chromatin OF DEAD AND DYING CELLS |
| CN100410651C (en) * | 2006-01-12 | 2008-08-13 | 上海交通大学 | Teaching kit for water contamination indication of horsebean micronucleus |
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2007
- 2007-09-17 CN CN2007101324493A patent/CN101131385B/en not_active Expired - Fee Related
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| RU2461894C1 (en) * | 2011-07-12 | 2012-09-20 | Государственное образовательное учреждение высшего профессионального образования "Северо-Осетинская государственная медицинская академия" Министерства здравоохранения и социального развития РФ | Method for prevention of toxic action of cadmium in experimental animals suffering chronic poisoning |
| RU2462764C1 (en) * | 2011-07-13 | 2012-09-27 | Государственное образовательное учреждение высшего профессионального образования "Северо-Осетинская государственная медицинская академия" Министерства здравоохранения и социального развития РФ | Method for prevention of toxic action of cadmium in experimental animals suffering chronic poisoning |
| RU2462763C1 (en) * | 2011-07-13 | 2012-09-27 | Государственное образовательное учреждение высшего профессионального образования "Северо-Осетинская государственная медицинская академия" Министерства здравоохранения и социального развития РФ | Method for prevention of toxic action of cadmium in experimental animals suffering chronic poisoning |
| CN102433371A (en) * | 2011-10-18 | 2012-05-02 | 南京师范大学 | Overall rotifer male fluorescent staining and in vitro sperm observation method |
| CN102433371B (en) * | 2011-10-18 | 2014-03-26 | 南京师范大学 | Overall rotifer male fluorescent staining and in vitro sperm observation method |
| CN105076108A (en) * | 2015-07-07 | 2015-11-25 | 安徽师范大学 | Short-term fixing preservation method and long-term fixing preservation method for rotifer specimen |
| CN105076108B (en) * | 2015-07-07 | 2018-05-01 | 安徽师范大学 | The short-term fixed store method and long-term fixed store method of wheel animalcule sample |
| CN108485981A (en) * | 2018-03-27 | 2018-09-04 | 中国科学院城市环境研究所 | A kind of separation of arenaceous shell worm and culture technique |
| CN108485981B (en) * | 2018-03-27 | 2020-12-08 | 中国科学院城市环境研究所 | Separation and culture technology of arenaria |
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| CN101131385B (en) | 2010-11-10 |
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Granted publication date: 20101110 Termination date: 20110917 |