CN110964721B - Simple and rapid grinding method for trace biological tissues - Google Patents
Simple and rapid grinding method for trace biological tissues Download PDFInfo
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- CN110964721B CN110964721B CN201911384732.4A CN201911384732A CN110964721B CN 110964721 B CN110964721 B CN 110964721B CN 201911384732 A CN201911384732 A CN 201911384732A CN 110964721 B CN110964721 B CN 110964721B
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- 238000000227 grinding Methods 0.000 title claims abstract description 76
- 238000000034 method Methods 0.000 title claims abstract description 42
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims abstract description 126
- 239000007788 liquid Substances 0.000 claims abstract description 63
- 229910052757 nitrogen Inorganic materials 0.000 claims abstract description 63
- 238000007710 freezing Methods 0.000 claims abstract description 50
- 230000008014 freezing Effects 0.000 claims abstract description 49
- 229910000831 Steel Inorganic materials 0.000 claims abstract description 27
- 239000010959 steel Substances 0.000 claims abstract description 27
- 230000001502 supplementing effect Effects 0.000 claims abstract description 5
- 239000000523 sample Substances 0.000 claims description 62
- 239000000463 material Substances 0.000 claims description 5
- 238000005498 polishing Methods 0.000 claims description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 3
- 239000012472 biological sample Substances 0.000 claims description 2
- 238000011049 filling Methods 0.000 claims description 2
- 230000002906 microbiologic effect Effects 0.000 claims description 2
- 102000039446 nucleic acids Human genes 0.000 abstract description 9
- 108020004707 nucleic acids Proteins 0.000 abstract description 9
- 150000007523 nucleic acids Chemical class 0.000 abstract description 9
- 102000004169 proteins and genes Human genes 0.000 abstract description 7
- 108091005461 Nucleic proteins Proteins 0.000 abstract description 6
- 238000012864 cross contamination Methods 0.000 abstract description 6
- 238000000605 extraction Methods 0.000 abstract description 5
- 241000233866 Fungi Species 0.000 abstract description 3
- 241001465754 Metazoa Species 0.000 abstract description 3
- 238000002474 experimental method Methods 0.000 abstract description 3
- 210000001519 tissue Anatomy 0.000 description 13
- 238000002360 preparation method Methods 0.000 description 9
- 241000209094 Oryza Species 0.000 description 8
- 235000007164 Oryza sativa Nutrition 0.000 description 8
- 235000009566 rice Nutrition 0.000 description 8
- 239000000843 powder Substances 0.000 description 6
- 238000005303 weighing Methods 0.000 description 6
- 230000000694 effects Effects 0.000 description 4
- 229920002521 macromolecule Polymers 0.000 description 3
- 241000196324 Embryophyta Species 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
- 238000004140 cleaning Methods 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- 230000000593 degrading effect Effects 0.000 description 2
- 238000007689 inspection Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000000751 protein extraction Methods 0.000 description 2
- 238000010257 thawing Methods 0.000 description 2
- 101710163270 Nuclease Proteins 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 230000002528 anti-freeze Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 238000005336 cracking Methods 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 230000002427 irreversible effect Effects 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 210000003491 skin Anatomy 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 210000001835 viscera Anatomy 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
- C12N15/1003—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/06—Lysis of microorganisms
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Abstract
The invention discloses a simple and rapid grinding method for trace biological tissues, which comprises the steps of placing samples into a centrifuge tube, adding 1-2 steel balls, covering the centrifuge tube, placing the centrifuge tube into a freezing paper box filled with liquid nitrogen, supplementing volatilized liquid nitrogen during the period, covering a cover of the freezing paper box after all the samples are added, freezing the samples for 0.5-1 min, pouring the liquid nitrogen into a liquid nitrogen tank, manually and rapidly shaking the freezing paper box up and down for 0.5-1 min to crush the samples, pouring the liquid nitrogen into the freezing paper box again to fill the freezing paper box, covering the freezing paper box cover, keeping the samples in the protection of the liquid nitrogen, and preventing the samples from being thawed and degraded, and opening the cover of the freezing paper box to take out the centrifuge tube when the samples are needed in experiments. The sample in the centrifuge tube can be directly used for the subsequent extraction of nucleic acids and proteins. The method has the advantages of small sample loss, simple and convenient operation and strong practicability, and can not only improve the efficiency, but also avoid the cross contamination of samples. Can be widely applied to grinding of plant, animal, fungi and other trace tissues.
Description
Technical Field
The invention relates to a simple and rapid grinding method for trace biological tissues, which can be widely applied to grinding of trace tissues such as plants, animals, fungi and the like, and belongs to the technical field of biochemical and molecular biological research.
Background
In the field of biochemical and molecular biological research, grinding of micro-biological tissues is mostly an essential stage in the preparation of biological macromolecules such as active proteins or nucleic acids. The grinding is a physical friction heat generating process, the activity of biological macromolecules is easy to be reduced or lost after long-time grinding, in order to prevent the activity from being reduced or lost, a liquid nitrogen grinding method is generally adopted in a laboratory for grinding trace biological tissues, and the tissues are frozen and crisp due to the fact that liquid nitrogen freezing can effectively inhibit the activity of nuclease and protease at ultralow temperature, and the grinding effect is best. The traditional grinding method is to put a sample into a centrifuge tube, freeze the centrifuge tube by liquid nitrogen, and then manually grind the sample by using a grinding rod. The traditional method can only grind one sample at a time, and for hundreds of samples, the efficiency is low, time and labor are wasted, operators always hold the centrifuge tube for a long time after grinding tens of samples, the left hand is in a frozen state, the right hand repeatedly rotates the grinding rod, and bubbles are easily generated, so that the hands of the operators are most easily damaged. The sample is easy to pollute and splash in the grinding process due to the open working state, so that the sample is lost, and meanwhile, in order to ensure that the sample is not thawed, the centrifuge tube is required to be frequently frozen by liquid nitrogen, so that time is wasted. Aiming at the traditional grinding method, some students reform the grinding rod or invent a new grinding device, but only one grinding device can be used at a time, the working efficiency can not be improved well, and the practicability is not strong. At present, some companies sell corresponding biological tissue grinding instruments, but the grinding instruments are expensive, and the grinding modes of many grinding instruments are easy to cause the breakage of a centrifuge tube, so that grinding failure is caused, and the grinding is irreversible loss for rare samples. In addition, although the samples can be frozen by liquid nitrogen before and after the machine is started, the liquid nitrogen cannot be added into the grinding instrument, each sample needs to be independently started and started, and for a large number of samples, the time required by the operation is long, the samples are easy to defrost, and the degradation of the samples is easy to cause.
Therefore, for the extraction of active biological macromolecules, it is necessary to develop a simple and rapid grinding method for trace biological tissues, reduce the loss degree of samples in grinding, reduce the cross contamination of samples and improve the grinding efficiency.
Disclosure of Invention
In order to solve the technical problems, the invention provides a simple and rapid grinding method for trace biological tissues, which can reduce the sample loss, is simple and convenient to operate, has strong practicability, can improve the efficiency, and can avoid cross contamination of samples.
The invention is realized by the following technical scheme:
a simple and rapid grinding method for trace biological tissues comprises the following steps:
(1) Preparing grinding consumable materials;
(2) Pouring liquid nitrogen into a freezing paper box, filling the freezing paper box with the liquid nitrogen without overflowing, adding biological samples to be ground into sterilized centrifuge tubes, putting 1-2 sterilized steel balls into each centrifuge tube, covering a centrifuge tube cover, putting the centrifuge tubes into a grid of the freezing paper box, supplementing the volatilized liquid nitrogen during the period to fill the freezing paper box with the liquid nitrogen without overflowing, and covering the freezing paper box with the cover of the freezing paper box after all the samples are added;
(3) After the sample is frozen for 0.5-1 min, pouring liquid nitrogen into the liquid nitrogen tank from one corner of the freezing paper box, and then covering the freezing paper box with a cover;
(4) Manually shaking the freezing paper box up and down at a speed of not less than 2 times per second to break the sample; the number of times per second is not less than 2, and the number of times is 1 from top to bottom in a manual continuous mode;
(5) Uncovering the cover of the frozen paper box, pouring liquid nitrogen into the frozen paper box again to fill the frozen paper box without overflowing, then covering the cover of the frozen paper box, so that the sample is in the protection of liquid nitrogen, the thawing degradation of the sample is prevented, the cover of the frozen paper box is opened when the sample is required for the experiment, the centrifuge tube is taken out, and the sample in the centrifuge tube is directly used for the subsequent extraction of nucleic acid and protein.
Further, the diameter of the steel ball in the step (2) is 0.5 to 0.7 times of the inner diameter of the centrifuge tube, for example, 0.5 times, 0.55 times, 0.6 times, 0.65 times and 0.7 times of the inner diameter of the centrifuge tube can be used.
Further, the total time for shaking the freezer paper tray in the step (4) is 0.5 to 1min, and may be, for example, 0.5min, 0.6min, 0.7min, 0.8min, 0.9min, 1min.
Further, in order to recycle the steel ball balls and prevent the sample from thawing and degrading, in the step (5), pouring liquid nitrogen into the freezing paper box again, taking out the centrifuge tube by using tweezers, pouring out the steel ball balls, leaving the sample in the centrifuge tube, covering the centrifuge tube cover, putting the centrifuge tube back into the grid of the freezing paper box, and covering the freezing paper box cover; cleaning the poured steel balls with clear water, sterilizing and drying for later use.
The grinding consumable in the step (1) comprises a plastic centrifuge tube, a steel ball, a freezing paper box, liquid nitrogen, anti-freezing gloves and tweezers.
And (3) repeating the step (3) and the step (4) for 1-2 times when the fresh sample amount is more than 120mg or the dry sample is more than 60mg so as to ensure that the sample is sufficiently ground, wherein the sample sufficiently ground means that the sample is ground into powder and can be used for subsequent extraction of nucleic acid and protein.
Compared with the prior art, the invention has the beneficial effects that:
1. the sample loss is small. The invention adopts totally-enclosed grinding, only a small amount of sample is lost when the steel ball is separated from the sample, if the operation is proper, the sample is not easy to be attached to the steel ball under the frozen state because the steel ball is arranged on the sample, and the sample is not easy to be taken out along with the steel ball when the steel ball is poured out. If the steel ball is not required to be recycled, the steel ball is not required to be poured out, so that the loss of the sample is almost zero. The method of the invention is particularly applicable to scarce samples.
2. The liquid nitrogen loss is low. In the operation process of the invention, the liquid nitrogen is not in direct contact with the sample, and can be recycled after the use, thereby obviously reducing the loss of the liquid nitrogen. Meanwhile, liquid nitrogen is in a relatively closed environment in the operation process, so that volatilization of the liquid nitrogen is reduced, and the utilization rate of the liquid nitrogen is improved.
3. The operation is safe and simple. The whole flow is simple, the grinding process of the traditional method is not needed, the working intensity is reduced, the operators do not need to have special skills, meanwhile, the direct contact step of the operators and liquid nitrogen is reduced in the operation process, the probability of personnel injury is greatly reduced, and the operation safety is improved.
4. The working efficiency is high. Through multiple operation statistics of different experimenters, the preparation time of grinding consumable materials and the sample weighing time are eliminated, and 100 samples are ground (the sample of less than or equal to 120 mg) by adopting the method, if the steel ball is not recovered, the time is 10-18 min, and if the steel ball is recovered, the time is 17-30 min. Compared with the traditional grinding method, the method can save more than 92 percent of time compared with the traditional grinding method for grinding 100 samples (the samples less than or equal to 120 mg), and has obvious advantages.
5. No cross contamination. The invention adopts totally-enclosed grinding, samples are absolutely independent in the whole grinding process, cross contamination can not exist, and the samples are isolated from liquid nitrogen, so that the possible contamination between the samples and the liquid nitrogen can be effectively avoided on the premise of ensuring the cleaning of the steel balls.
6. Can effectively prevent the centrifuge tube from cracking and the sample from degrading. Because most of the commercial grinding instruments at present have larger grinding force and are easy to cause the breakage of the centrifuge tube, the invention adopts manual control to shake the freezing paper box, thereby effectively preventing the breakage of the centrifuge tube. In addition, the liquid nitrogen cannot be added into the grinding instrument after the sample is put on the machine by adopting the grinding instrument, and each sample needs to be put on the machine and put off the machine independently.
7. The invention is suitable for grinding and crushing various plant tissues (such as roots, stems, leaves, flowers, fruits and the like), animal tissues (such as muscles, internal organs, skin and the like), fungi and other trace fresh samples or dry samples, and has wide application range.
Detailed Description
The invention provides a simple and rapid grinding method for trace biological tissues, and a person skilled in the art can properly improve the process parameters by referring to the content of the invention. It is expressly noted that all such similar substitutions and modifications will be apparent to those skilled in the art, and are deemed to be included within the present invention. The invention will be further illustrated with reference to specific examples.
Example 1
100 fresh rice leaves were ground, 120mg per sample.
(1) Preparation of the grinding consumable. Comprises a plastic centrifuge tube with the volume of 2mL and the inner diameter of 10mm, a steel ball with the diameter of 6mm, a freezing paper box with the size of 100 grids (the height of 51mm is 158mm is wide), liquid nitrogen, antifreezing gloves and tweezers. These polishing consumables are commercially available.
(2) Liquid nitrogen is poured into the freezer cartons so that the cartons are filled with liquid nitrogen and do not overflow. 120mg of fresh rice leaf samples are added into each sterilized centrifuge tube, 1 ball of sterilized steel balls is placed into each centrifuge tube, the caps of the centrifuge tubes are covered, and the centrifuge tubes are placed into the grids of the freezing paper boxes. The liquid nitrogen was readily volatilized, during which time the liquid nitrogen was replenished 2 times to fill the freezer carton with liquid nitrogen without spilling until 100 samples were added, and the freezer carton was capped with the freezer carton lid.
(3) After the sample was frozen for 1min, the antifreeze glove was put on, the freezer paper box was carefully flattened, liquid nitrogen was poured back into the liquid nitrogen tank from one corner of the freezer paper box, and the freezer paper box lid was closed.
(4) After the liquid nitrogen is completely poured, the freezing paper box is rapidly shaken up and down manually at a speed of not less than 2 times per second, wherein the speed of not less than 2 times per second is 1 time manually and continuously; after 1min, the shaking was stopped, the lid of the freezer was opened, liquid nitrogen was poured into the freezer again to fill the freezer without spilling, and the samples were checked for complete grinding (powder).
(5) Taking out each centrifuge tube respectively by using tweezers, opening the cover of the centrifuge tube, carefully pouring out the ball of the steel ball, leaving the sample in the centrifuge tube, covering the cover of the centrifuge tube, putting the centrifuge tube back into the grid of the freezing paper box, and supplementing liquid nitrogen for 1 time during which the freezing paper box is filled with liquid nitrogen and does not overflow, and covering the cover of the freezing paper box, so that the sample is in the protection of the liquid nitrogen. The poured steel balls are cleaned by clear water, sterilized and dried for standby.
In the embodiment 1, through multiple operation statistics of different experimenters, the preparation time of grinding consumable materials and the weighing time of samples are eliminated, 100 samples are ground, the time is 17-30 min, all the samples are ground into powder after inspection, and the method can be directly used for extracting nucleic acid and protein.
Example 2
This example 2 was identical to example 1, except that the operation of removing the ball was not performed in step (5).
In the embodiment 2, through different experimenters and multiple operation statistics, the preparation time of grinding consumable materials and the weighing time of samples are eliminated, 100 samples are ground (steel balls are not recovered), the time is 10-18 min, and all the samples are ground into powder after inspection, so that the method can be directly used for extracting nucleic acid and protein.
Example 3
100 parts of dry rice leaves were ground, 60mg per sample.
The procedure and conditions were the same as in example 1, except that the following procedure and conditions were different in example 3.
The sample added in step (1) was a 60mg dry rice leaf sample.
The time for freezing the sample in step (3) was 0.5min.
The time for shaking the freezer paper box in step (4) was 0.5min.
In this example 3, the preparation time of the grinding consumable and the weighing time of the sample were excluded, 100 samples (dry sample, recovered ball) were ground, the time required was 16-29 min, and all samples were checked to be ground into powder, which was used directly for nucleic acid and protein extraction.
Example 4
Grinding 100 fresh rice leaves, 150mg (greater than 120mg sample) per sample
The procedure and conditions were the same as in example 1, except that the following procedure and conditions were different in example 4.
150mg of fresh rice leaf sample was added to each sterilized centrifuge tube in step (2).
Repeating the step (3) and the step (4) once.
In this example 4, the preparation time of the grinding consumable and the sample weighing time were excluded, 100 samples (fresh large sample, recovered ball) were ground, the time required was 19-32 min, and all samples were checked to be ground into powder, which could be used directly for nucleic acid and protein extraction.
Comparative example 1
The traditional grinding method is adopted to grind fresh rice leaf samples, and the specific operation is as follows:
(1) Preparation of the grinding consumable. Comprises a plastic centrifuge tube with the volume of 2mL and the inner diameter of 10mm, a centrifuge tube double-sided plate, a freezing paper box with 100 grids (the height of 51mm is 158mm, the width of 158 mm), a grinding rod, liquid nitrogen and antifreezing gloves. These polishing consumables are commercially available.
(2) Liquid nitrogen is poured into the freezing paper box, so that the freezing paper box is filled with the liquid nitrogen and does not overflow. Taking 1 sterilized centrifuge tube, adding 120mg of fresh rice leaf sample, taking out sterilized grinding rod by forceps, putting the grinding rod into the centrifuge tube, putting the centrifuge tube into a freezing paper box filled with liquid nitrogen, taking out the centrifuge tube after the sample is frozen for 0.5-1 min, putting the centrifuge tube into a double-sided board of the centrifuge tube, rapidly rotating the grinding rod, and supplementing the volatilized liquid nitrogen for 5-7 times during the period.
(3) And (3) putting the centrifuge tube into a freezing paper box filled with liquid nitrogen again, taking out the centrifuge tube, putting the centrifuge tube into a double-sided board of the centrifuge tube after the sample is frozen for 0.5-1 min, and rapidly rotating the grinding rod.
(4) Taking away the grinding rod, covering the cover of the centrifuge tube, putting the centrifuge tube into a freezing paper box filled with liquid nitrogen, and finishing grinding of a sample after the subsequent extraction of nucleic acid and protein.
Grinding other samples, and repeating the steps (2) to (4).
Through statistics, the time for grinding one sample is 2-4 min, 100 samples are ground, the preparation time and the sample weighing time of experimental consumables are eliminated, the time is 200-400 min, moreover, experimental operators are frozen by the liquid nitrogen for a long time in direct contact with a centrifuge tube, the left hand is frozen to be red, the right hand repeatedly rotates a grinding rod, the grinding rod is ground to form bubbles, and the follow-up experiment finds that part of samples are polluted, and a plurality of broken blades are in the residual liquid nitrogen and cannot be recycled.
As can be seen from the above examples and comparative examples, the method of the present invention has the characteristics of small sample loss, simple and convenient operation, strong practicality, capability of avoiding sample cross contamination, high working efficiency, and outstanding technical advantages compared with the conventional grinding method.
The foregoing is merely a preferred embodiment of the present invention and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present invention, which are intended to be comprehended within the scope of the present invention.
Claims (6)
1. The simple and rapid grinding method for the trace biological tissues is characterized by comprising the following steps of:
(1) Preparing grinding consumable materials;
(2) Pouring liquid nitrogen into a freezing paper box, filling the freezing paper box with the liquid nitrogen without overflowing, adding biological samples to be ground into sterilized centrifuge tubes, putting 1-2 sterilized steel balls into each centrifuge tube, covering a centrifuge tube cover, putting the centrifuge tubes into a grid of the freezing paper box, supplementing the volatilized liquid nitrogen during the period to fill the freezing paper box with the liquid nitrogen without overflowing, and covering the freezing paper box cover after all the samples are added;
(3) After the sample is frozen for 0.5-1 min, pouring liquid nitrogen into a liquid nitrogen tank from one corner of the freezing paper box, and then covering the freezing paper box with a cover;
(4) Manually shaking the freezing paper box up and down at a speed of not less than 2 times per second to break the sample; the number of times per second is not less than 2, and the number of times is 1 from top to bottom in a manual continuous mode;
(5) The lid of the freezer carton is opened, liquid nitrogen is poured into the freezer carton again, the freezer carton is filled without spilling, and then the lid of the freezer carton is covered.
2. The method for simply and rapidly grinding a trace biological tissue according to claim 1, wherein the diameter of the steel ball in the step (2) is 0.5-0.7 times of the inner diameter of a centrifuge tube used.
3. The method for simple and rapid polishing of a trace amount of biological tissue according to claim 1, wherein the total time for shaking the tissue box in the step (4) is 0.5 to 1min.
4. The simple and rapid grinding method for trace biological tissues according to claim 1, wherein in the step (5), after liquid nitrogen is poured into a freezing paper box again, a centrifuge tube is taken out by forceps, steel balls are poured out, a sample is left in the centrifuge tube, a cover of the centrifuge tube is covered, the centrifuge tube is put back into a grid of the freezing paper box, then the cover of the freezing paper box is covered, and the poured steel balls are washed with clean water, sterilized and dried for standby.
5. The method for simple and rapid grinding of micro-biological tissues according to claim 1, wherein the grinding consumable in the step (1) comprises a plastic centrifuge tube, a steel ball, a freezing paper box, liquid nitrogen, freezing gloves and tweezers.
6. The method according to any one of claims 1 to 5, wherein the steps (3) to (4) are repeated 1 to 2 times when the amount of the ground fresh sample is more than 120mg or the dry sample is more than 60mg.
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| CN105018473A (en) * | 2015-09-02 | 2015-11-04 | 中国科学院新疆生态与地理研究所 | Method for crushing samples in batches for plant RNA extraction |
| CN108913648A (en) * | 2018-07-12 | 2018-11-30 | 福建农林大学 | A kind of extracting method and its application of fresh leaves of tea plant fine hair |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105018473A (en) * | 2015-09-02 | 2015-11-04 | 中国科学院新疆生态与地理研究所 | Method for crushing samples in batches for plant RNA extraction |
| CN108913648A (en) * | 2018-07-12 | 2018-11-30 | 福建农林大学 | A kind of extracting method and its application of fresh leaves of tea plant fine hair |
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| Title |
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| J.A.Sweigard et al..A miniprep procedure for isolating genomic DNA from Magnaporthe grisea.《Fungal genetics reports》.1990,第37卷(第37期),第1-4页. * |
| 熊福全等.液氮研磨对桉树木粉颗粒结构的影响.《南京林业大学学报（自然科学版）》.2014,第38卷(第38期),第125-129页. * |
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