CN102250875A - Method for extracting RNAs from oleaginous microorganisms - Google Patents
Method for extracting RNAs from oleaginous microorganisms Download PDFInfo
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- CN102250875A CN102250875A CN 201010176579 CN201010176579A CN102250875A CN 102250875 A CN102250875 A CN 102250875A CN 201010176579 CN201010176579 CN 201010176579 CN 201010176579 A CN201010176579 A CN 201010176579A CN 102250875 A CN102250875 A CN 102250875A
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Abstract
The invention discloses a method for extracting RNAs from oleaginous microorganisms, which comprises the following steps: performing cell breakage of the oleaginous microorganisms by combining liquid nitrogen grinding and glass bead oscillation; adding an organic solvent into cell breakage solution to remove fat; adding acidic phenol/chloroform/isoamylol to remove protein; centrifuging, collecting a water phase, adding isopropanol and precipitating; and washing and precipitating by using ethanol, drying under vacuum, and successfully extracting the RNAs from the oleaginous microorganisms. The method is simple in operation and low in cost and ensures high integrity of the RNAs. The RNAs obtained by the method can meet the requirements for study on oleaginous microorganism in molecular biological aspects, such as Northern hybridization, mRNA separation, rapid amplification of cDNA ends (RACE), quantitative polymerase chain reaction (PCR), cDNA synthesis and in-vitro translation.
Description
Technical field
The present invention relates to the extracting method of a kind of microorganism RNA, particularly relate to the extracting method of oleaginous microorganism RNA.
Background technology
Occurring in nature part microorganism as bacterium, yeast, mould and algae etc., can store the grease that quality surpasses its dry cell weight 20% (w/w) in born of the same parents under specific culture condition, the microorganism with this phenotype is called as oleaginous microorganism.The grease that these oleaginous microorganisms produced mainly exists with the form of fatty acid triglycercide, and its composition is similar to general animal-plant oil.Microbe oil fermentation also becomes the important research direction of biofuel industry and industrial biotechnology gradually in recent years.With respect to general microorganism, fat content is higher in the oleaginous microorganism born of the same parents.Bacterial strain as some Rhodotorula can be the substratum fermentation of carbon source with the saccharine material, the grease (Li YH, Zhao ZB, the Bai FW.Enzyme Microb Technol that surpass its dry cell weight 70% (w/w) at intracellular accumulation, 2007 (41): 312-317), and also a large amount of simultaneously synthetic colour.These high-load greases and pigment produce very big interference to the separation and purification meeting of RNA, have a strong impact on quality and the output of extracting RNA.And the high-quality RNA of separation and Extraction is the basis of researchs such as genetic expression, regulation and control and genetically engineered.Microorganism RNA extracting method has much at present, adopt Trizol test kit (Gilchrist M separately, MacDonald AJ, Neverova I, et al.J Immunol methods, 1997 (201): 207-214), hot acid phenol is handled (Schmitt ME, Brown, TA, Trumpower LB.Nucleic Acids Res, 1990 (18): 3091-3092), granulated glass sphere smudge cells (Ao Sibai F, James Kingston R, Sai Deman J. fine works molecular biology experiment guide .1998:597-598) and liquid nitrogen grinding (Nivens DE.Curr Opin Biotechnol.2001 (12): 455-60) etc. method all can not effectively be extracted high-quality RNA from oleaginous microorganism for Sayler GS, Fleming JT.
Summary of the invention
The object of the present invention is to provide a kind of economy, simple operation method, from oleaginous microorganism, extract the cell total rna of high quality (high purity).
To achieve these goals, operation steps of the present invention is:
A kind of method of extracting high quality RNA from oleaginous microorganism, produce oil thalline at first adopt the liquid nitrogen grinding powdered, adopt granulated glass sphere vibration (mode that combines) to carry out cytoclasis in RNA extraction damping fluid then, get cytoclasis liquid; In cytoclasis liquid, add organic solvent and remove grease, centrifugal back water intaking phase adds sodium-acetate, acid phenol, chloroform and primary isoamyl alcohol and removes intracellular protein, and the water intaking of centrifugal back is added to isopropanol precipitating RNA, washing precipitation adds the dissolving of DEPC treating water after the vacuum-drying.
Specific operation process is,
1) getting cell density is 1 * 10
7-1 * 10
8The thalline of cell/ml grinds broken 0.5-30min to Powdered through the produce oil thalline of liquid nitrogen flash freezer, scrapes to get powder and be loaded in the centrifuge tube, adds RNA and extracts damping fluid, and thalline is 10ml: 1-2ml with RNA extraction damping fluid volume ratio;
2) add the granulated glass sphere interrupted oscillation in centrifuge tube, total broken time is 0.5-30min; Every 10ml thalline (in the 5ml centrifuge tube) adds the 0.5-1.5g granulated glass sphere;
3) in centrifuge tube, add organic solvent, fully behind the mixing 0-4 ℃, the centrifugal 5-10min of 12000-18000 * g; The amount of organic solvent is that 1/4-3/4RNA extracts the damping fluid volume;
4) water of getting after the layering is transferred in the clean centrifuge tube, and adding concentration is that 3M, pH are that 4.6 sodium-acetate and volume ratio are 25: 24: 1 acid phenol/chloroform/primary isoamyl alcohol mixed solutions, and fully mixing 0-4 ℃, the centrifugal 5-15min of 12000-18000 * g; By the 10ml thalline, add sodium-acetate and the 1ml mixed solution of 100 μ l;
5) water of getting after the layering is transferred in the clean centrifuge tube, adds equal-volume Virahol mixing, places 0--70 ℃ to place 5min-16h, and 0-4 ℃ then, the centrifugal 5-15min of 12000-18000 * g obtains sedimentary RNA;
6) the RNA precipitation that obtains is preserved with the DEPC treating water dissolving that adds volume ratio 0.01%-0.5% after 70% the washing with alcohol drying.
It is the aqueous solution that includes 4M guanidinium isothiocyanate, 0.5wt% sarcosyl, 25mM Trisodium Citrate, pH 7.0,0.5wt% mercaptoethanol that RNA extracts damping fluid.
Described granulated glass sphere vibration breaking method is an interrupted oscillation, and the 10s-10min that at every turn vibrates is placed on cooled on ice 0.5-10min, and total broken time is 0.5-30min, and the granulated glass sphere diameter is 0.2-1mm.
The greasy organic solvent of described removal is that chloroform, methylene dichloride, sherwood oil or the arbitrary proportion between them mix.
The present invention is different from traditional microorganism RNA extracting method, for smudge cells fast, adopts liquid nitrogen grinding and granulated glass sphere oscillating phase bonded mode.
The present invention adds organic reagent, has effectively removed a large amount of greases and pigment in the oleaginous microorganism born of the same parents, has reduced the interference to the RNA separation and purification.
The oleaginous microorganism that the present invention uses includes but not limited to the produce oil fungi, as Rhodosporidiumtoruloides, Cryptococcus curvatus, Yarrowia lipolytica, Rhodotorula glutinis, Rhizopus arrhizus, Lipomyces starkeyi, Mortierella isabellina, Mucorcircinelloides and Cunninghamella; The produce oil bacterium is as Croynebacterium, Nocardia and Mycobacterium; The little algae of produce oil is as Botryococcus braunii, Crypthecodiniumcohnii, Chlorella protothecoides, Nannochloropsis sp. and Schizochytriumlimacinum.The high quality RNA of gained can be widely used in as Northern hybridization, mRNA separation, RACE, quantitative PCR, cDNA synthetic and external translation etc.
Description of drawings
Fig. 1. oleaginous yeast Lipomyces starkeyi AS 2.1560 total RNA extract electrophoretic analysis, swimming lane 1: do not add organic reagent and remove grease; 2: add organic reagent and remove grease (following identical).
Fig. 2. oleaginous yeast Rhodosporidium toruloides AS 2.1389 total RNA extract electrophoretic analysis.
Fig. 3. the total RNA of the little algae Botryococcus of produce oil braunii LB572 extracts electrophoretic analysis.
Fig. 4. oleaginous yeast Yarrowia lipolytica AS 2.1398 total RNA extract electrophoretic analysis.
Fig. 5. produce oil mould Mucor circinelloides AS 3.2208 total RNA extract electrophoretic analysis.
Fig. 6. oleaginous yeast Rhodotorula glutinis AS 2.703 total RNA extract electrophoretic analysis.
Fig. 7. the total RNA of produce oil bacterium Corynebacterium glutanicum extracts electrophoretic analysis.
Embodiment
Following examples have been chosen some typical oleaginous microorganism bacterial strains, and the extracting method of oleaginous microorganism RNA has been described, this will help to understand this patent, not use the present invention but be not limited in any form on other bacterial strain material.Agents useful for same among the following embodiment:
1.RNA extraction damping fluid: 4M guanidinium isothiocyanate, 0.5% sarcosyl, 25mM Trisodium Citrate, pH 7.0,0.5% mercaptoethanols;
2.DEPC treating water: the 100ml deionized water adds 100 μ l DEPC, mixing under the normal temperature, autoclaving, the DEPC treating water of volume ratio 0.1%;
3. sodium-acetate: the 24.61g sodium-acetate is dissolved in the DEPC treating water, and regulating pH with Glacial acetic acid is 4.6, and constant volume is put 100ml, autoclaving.
Embodiment one: it is 1 * 10 that oleaginous yeast Lipomyces starkeyi AS 2.1560 (bacterium source in Chinese common micro-organisms culture presevation administrative center) is cultivated cell density
8Cell/ml collects 10ml thalline liquid nitrogen flash freezer.Yeast cell liquid nitrogen grinding 10min is to Powdered, scrape and get powder and be loaded in the 5ml centrifuge tube, the RNA that adds the 1ml precooling extracts damping fluid, mixing, adding the 1.5g diameter is the granulated glass sphere of 0.4mm, vortex vibration 10min, 10min intermittently, total broken time is 30min, adds behind the 500 μ l chloroforms fully mixing (control group does not add chloroform), 13, the centrifugal 10min of 000 * g, carefully draw water, add the sodium-acetate of 100 μ l successively, acid phenol/chloroform/the primary isoamyl alcohol (volume ratio is 25: 24: 1) of 1ml is put upside down commentaries on classics and is mixed several times, ice bath 15min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions is transferred to water in the clean centrifuge tube, and add isopyknic Virahol, mixing is placed on-70 ℃ and places 1h, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions, remove supernatant liquor, 70% ethanol is washed precipitation once, adds 100 μ l DEPC treating water precipitation after the vacuum-drying, places under-70 ℃ of cold condition and preserves.
Embodiment two: it is 1 * 10 that oleaginous yeast Rhodosporidium toruloides AS 2.1389 (bacterium source in Chinese common micro-organisms culture presevation administrative center) is cultivated cell density
8Cell/ml collects 10ml thalline liquid nitrogen flash freezer.Yeast cell liquid nitrogen grinding 30min is to Powdered, scrape and get powder and be loaded in the 5ml centrifuge tube, the RNA that adds the 1ml precooling extracts damping fluid, mixing, adding the 1.5g diameter is the granulated glass sphere of 0.4mm, vortex vibration 0.5min, 0.5min intermittently, total broken time is 1min, adds behind the 500 μ l chloroforms fully mixing (control group does not add chloroform), 13, the centrifugal 10min of 000 * g, carefully draw water, add the sodium-acetate of 100 μ l successively, acid phenol/chloroform/the primary isoamyl alcohol (volume ratio is 25: 24: 1) of 1ml is put upside down commentaries on classics and is mixed several times, ice bath 15min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions is transferred to water in the clean centrifuge tube, and add isopyknic Virahol, mixing is placed on-20 ℃ and places 5min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions, remove supernatant liquor, 70% ethanol is washed precipitation once, adds 100 μ l DEPC treating water dissolution precipitations after the vacuum-drying, places under-70 ℃ of cold condition and preserves.
Embodiment three: it is 1 * 10 that the little algae Botryococcus of produce oil braunii LB572 (bacterium source is in Texas ,Usa university) is cultivated cell density
7Cell/ml collects 10ml thalline liquid nitrogen flash freezer.Cell liquid nitrogen grinding 2min is to Powdered, scrape and get powder and be loaded in the 5ml centrifuge tube, the RNA that adds the 1ml precooling extracts damping fluid, mixing, adding the 1.5g diameter is the 0.6mm granulated glass sphere, vortex vibration 3min, 3min intermittently, total broken time is 9min, adds behind the 500 μ l methylene dichloride fully mixing (control group does not add methylene chloride), 13, the centrifugal 10min of 000 * g, carefully draw water, add the sodium-acetate of 100 μ l successively, acid phenol/chloroform/the primary isoamyl alcohol (volume ratio is 25: 24: 1) of 1ml is put upside down commentaries on classics and is mixed several times, ice bath 15min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions is transferred to water in the clean centrifuge tube, and add isopyknic Virahol, mixing is placed on-50 ℃ and places 20min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions, remove supernatant liquor, 70% ethanol is washed precipitation once, adds 100 μ l DEPC treating water dissolution precipitations after the vacuum-drying, places under-70 ℃ of cold condition and preserves.
Embodiment four: it is 1 * 10 that oleaginous yeast Yarrowia lipolytica AS 2.1398 (bacterium source in Chinese common micro-organisms culture presevation administrative center) is cultivated cell density
8Cell/ml collects the 10ml thalline, liquid nitrogen flash freezer.Yeast cell liquid nitrogen grinding 10min is to Powdered, scrape and get powder and be loaded in the 5ml centrifuge tube, the RNA that adds the 1ml precooling extracts damping fluid, mixing, adding the 1.5g diameter is the granulated glass sphere of 0.4mm, vortex vibration 2min, 2min intermittently, total broken time is 6min, adds fully mixing (control group does not add chloroform/methylene dichloride) of 500 μ l chloroform/methylene dichloride (volume ratio is 1: 1) back, 13, the centrifugal 10min of 000 * g, carefully draw water, add the sodium-acetate of 100 μ l successively, acid phenol/chloroform/the primary isoamyl alcohol (volume ratio is 25: 24: 1) of 1ml is put upside down commentaries on classics and is mixed several times, ice bath 15min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions is transferred to water in the clean centrifuge tube, and add isopyknic Virahol, mixing is placed on-60 ℃ and places 16h, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions, the careful supernatant liquor of removing, 70% ethanol is washed precipitation once, adds 100 μ l DEPC treating water dissolution precipitations after the vacuum-drying, places under-70 ℃ of cold condition and preserves.
Embodiment five: it is 1 * 10 that produce oil mould Mucor circinelloides AS 3.2208 (bacterium source in Chinese common micro-organisms culture presevation administrative center) is cultivated cell density
8Cell/ml collects the 10ml thalline, liquid nitrogen flash freezer.Yeast cell grinds 15min to Powdered, scrape and get powder and be loaded in the 5ml centrifuge tube, the RNA that adds the 1ml precooling extracts damping fluid, mixing, adding the 1.5g diameter is the granulated glass sphere of 1mm, vortex vibration 5min, 5min intermittently, total broken time is 10min, adds behind the 500 μ l methylene dichloride fully mixing (control group does not add methylene chloride), 13, the centrifugal 10min of 000 * g, carefully draw water, add the sodium-acetate of 100 μ l successively, acid phenol/chloroform/the primary isoamyl alcohol (volume ratio is 25: 24: 1) of 1ml is put upside down commentaries on classics and is mixed several times, ice bath 15min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions is transferred to water in the clean centrifuge tube, and add isopyknic Virahol, mixing is placed on-20 ℃ and places 10h, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions, remove supernatant liquor, 70% ethanol is washed precipitation once, adds 100 μ l DEPC treating water dissolution precipitations after the vacuum-drying, places under-70 ℃ of cold condition and preserves.
Embodiment six: it is 1 * 10 that oleaginous yeast Rhodotorula glutinis AS 2.703 (bacterium source in Chinese common micro-organisms culture presevation administrative center) is cultivated cell density
8Cell/ml collects the 10ml thalline, liquid nitrogen flash freezer.Yeast cell liquid nitrogen grinding 10min is to Powdered, scrape and get powder and be loaded in the 5ml centrifuge tube, the RNA that adds the 1ml precooling extracts damping fluid, mixing, adding the 1g diameter is the granulated glass sphere of 0.2mm, vortex vibration 3min, 4min intermittently, total broken time is 15min, adds behind the 500 μ l sherwood oils fully mixing (control group does not add sherwood oil), 13, the centrifugal 5min of 000 * g, carefully get supernatant, add the sodium-acetate of 100 μ l successively, acid phenol/chloroform/the primary isoamyl alcohol (volume ratio is 25: 24: 1) of 1ml is put upside down commentaries on classics and is mixed several times, ice bath 15min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions, with the upper water phase transition to the clean centrifuge tube, and add isopyknic Virahol, mixing is placed on 0 ℃ and places 2h, the centrifugal 20min of 13,000 * g under 4 ℃ of conditions, the careful supernatant liquor of removing, 70% ethanol is washed precipitation once, adds 100 μ l DEPC treating water dissolution precipitations after the vacuum-drying, places under-70 ℃ of cold condition and preserves.
Embodiment seven: it is 1 * 10 that produce oil bacterium Corynebacterium glutanicum (bacterium source in Chinese common micro-organisms culture presevation administrative center) is cultivated cell density
8Cell/ml collects the 10ml thalline, liquid nitrogen flash freezer.Yeast cell liquid nitrogen grinding 0.5min is to Powdered, scrape and get powder and be loaded in the 5ml centrifuge tube, the RNA that adds the 1ml precooling extracts damping fluid, mixing, adding the 0.5g diameter is the granulated glass sphere of 0.2mm, vortex vibration 10s, 0.5min intermittently, total broken time is 0.5min, adds fully mixing (control group does not add sherwood oil/chloroform) of 500 μ l sherwood oil/chloroforms (volume ratio is 1: 1) back, 13, the centrifugal 10min of 000 * g, carefully draw water, add the sodium-acetate of 100 μ l successively, acid phenol/chloroform/the primary isoamyl alcohol (volume ratio is 25: 24: 1) of 1ml is put upside down commentaries on classics and is mixed several times, ice bath 15min, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions is transferred to water in the clean centrifuge tube, and add isopyknic Virahol, mixing is placed on-30 ℃ and places 2h, the centrifugal 15min of 13,000 * g under 4 ℃ of conditions, the careful supernatant liquor of removing, 70% ethanol is washed precipitation once, adds 100 μ l DEPC treating water dissolution precipitations after the vacuum-drying, places under-70 ℃ of cold condition and preserves.
What above embodiment explained, add or do not add the cell total rna that the interior grease of organic removal of solvents born of the same parents is extracted, their electrophoretic analysis result sees Figure of description 1-7 respectively.As can be seen from the figure, the cell total rna that adopts the inventive method to extract, electrophoresis detection 28S and 18S rRNA band are clear with respect to the method that does not add organic solvent, illustrate that RNA has kept good integrity.
Get and add the resulting RNA sample of organic reagent removal grease among the embodiment 1-7,50 times of adding distil water dilutions, and do blank with distilled water, the reading of regulating ultraviolet spectrophotometer at 230nm, 260nm, 280nm place is read testing sample in the OD at three wavelength places value then to zero respectively.Table 1 result shows: by the OD of the total RNA of UV spectrophotometer measuring
260/ OD
280Value all between 1.8-2.2, illustrates that RNA purity meets the requirements.
Table 1RNA purity detecting
Claims (8)
1. method of extracting high quality RNA from oleaginous microorganism is characterized in that:
The produce oil thalline at first adopts the liquid nitrogen grinding powdered, extracts employing granulated glass sphere vibration carrying out cytoclasis in the damping fluid in RNA then, gets cytoclasis liquid; In cytoclasis liquid, add organic solvent and remove grease, centrifugal back water intaking phase adds sodium-acetate, acid phenol, chloroform and primary isoamyl alcohol and removes intracellular protein, and the water intaking of centrifugal back is added to isopropanol precipitating RNA, washing precipitation adds the dissolving of DEPC treating water after the vacuum-drying.
2. it is characterized in that in accordance with the method for claim 1: described oleaginous microorganism is produce oil fungi, produce oil bacterium and/or the little algae of produce oil.
3. in accordance with the method for claim 2, it is characterized in that: described produce oil fungi is one or more among Rhodosporidium toruloides, Cryptococcus curvatus, Yarrowia lipolyica, Rhodotorula glutinis, Rhizopus arrhizus, Lipomyces starkeyi, Mortierellaisabellina, Mucor circinelloides, the Cunninghamella, and the produce oil bacterium is one or more among Croynebacterium, Nocardia and the Mycobacterium; The little algae of produce oil is one or more among Botryococcus braunii, Crypthecodinium cohnii, Chlorellaprotothecoides, Nannochloropsis sp. and the Schizochytrium limacinum.
4. in accordance with the method for claim 1, it is characterized in that: specific operation process is,
1) getting cell density is 1 * 10
7-1 * 10
8The thalline of cell/ml grinds broken 0.5-30min to Powdered through the produce oil thalline of liquid nitrogen flash freezer, scrapes to get powder and be loaded in the centrifuge tube, adds RNA and extracts damping fluid, and thalline is 10ml: 1-2ml with RNA extraction damping fluid volume ratio;
2) add the granulated glass sphere interrupted oscillation in centrifuge tube, total broken time is 0.5-30min; Every 10ml thalline adds the 0.5-1.5g granulated glass sphere;
3) in centrifuge tube, add organic solvent, fully behind the mixing 0-4 ℃, the centrifugal 5-10min of 12000-18000 * g; The amount of organic solvent is that 1/4-3/4RNA extracts the damping fluid volume;
4) water of getting after the layering is transferred in the clean centrifuge tube, and adding concentration is that 3M, pH are that 4.6 sodium-acetate and volume ratio are 25: 24: 1 acid phenol/chloroform/primary isoamyl alcohol mixed solutions, and fully mixing 0-4 ℃, the centrifugal 5-15min of 12000-18000 * g; By the 10ml thalline, add sodium-acetate and the 1ml mixed solution of 100 μ 1;
5) water of getting after the layering is transferred in the clean centrifuge tube, adds equal-volume Virahol mixing, places 0--70 ℃ to place 5min-16h, and 0-4 ℃ then, the centrifugal 5-15min of 12000-18000 * g obtains sedimentary RNA.
5. it is characterized in that in accordance with the method for claim 4: the RNA precipitation of acquisition is preserved with adding the dissolving of DEPC treating water after 70% the washing with alcohol drying.
6. according to claim 1 or 4 described methods, it is characterized in that: it is the aqueous solution that includes 4M guanidinium isothiocyanate, 0.5wt% sarcosyl, 25mM Trisodium Citrate, pH7.0,0.5wt% mercaptoethanol that RNA extracts damping fluid.
7. in accordance with the method for claim 4, its feature also is: described granulated glass sphere vibration breaking method is an interrupted oscillation, each vibration 10s-10min is placed on cooled on ice 0.5-10min, and total broken time is 0.5-30min, and the granulated glass sphere diameter is 0.2-1mm.
8. according to claim 1 or 4 described methods, its feature also is: the greasy organic solvent of described removal is that chloroform, methylene dichloride, sherwood oil or the arbitrary proportion between them mix.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102586233A (en) * | 2012-03-09 | 2012-07-18 | 江南大学 | Method for rapidly extracting total RNA (ribonucleic acid) in yeast |
| CN105779442A (en) * | 2016-05-19 | 2016-07-20 | 清华大学 | Extraction method of macro-transcriptome RNA of flora utilized by natural cellulose |
| CN105821031A (en) * | 2016-01-18 | 2016-08-03 | 昆明医科大学 | Principle of extracting DNA by using dichloromethane method |
| CN108048451A (en) * | 2017-12-26 | 2018-05-18 | 浙江海洋大学 | A kind of non-damaging live body cuttlefish DNA sample acquisition method and extracts reagent |
| CN109652407A (en) * | 2019-01-31 | 2019-04-19 | 华南理工大学 | A kind of extracting method of microalgae cell total serum IgE |
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| WO2008002740A2 (en) * | 2006-06-28 | 2008-01-03 | Sigma-Aldrich Co. | Methods, compositions, and kits for rna extraction |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2008002740A2 (en) * | 2006-06-28 | 2008-01-03 | Sigma-Aldrich Co. | Methods, compositions, and kits for rna extraction |
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| 《吉林农业科学》 20091231 邢晋祎等 鸡脂肪组织总RNA提取方法的优化和RT-PCR检测 41-44 1-8 第34卷, 第5期 * |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102586233A (en) * | 2012-03-09 | 2012-07-18 | 江南大学 | Method for rapidly extracting total RNA (ribonucleic acid) in yeast |
| CN102586233B (en) * | 2012-03-09 | 2013-07-10 | 江南大学 | Method for rapidly extracting total RNA (ribonucleic acid) in yeast |
| CN105821031A (en) * | 2016-01-18 | 2016-08-03 | 昆明医科大学 | Principle of extracting DNA by using dichloromethane method |
| CN105779442A (en) * | 2016-05-19 | 2016-07-20 | 清华大学 | Extraction method of macro-transcriptome RNA of flora utilized by natural cellulose |
| CN105779442B (en) * | 2016-05-19 | 2019-05-28 | 清华大学 | A kind of native cellulose using the macro transcript profile RNA of flora extracting method |
| CN108048451A (en) * | 2017-12-26 | 2018-05-18 | 浙江海洋大学 | A kind of non-damaging live body cuttlefish DNA sample acquisition method and extracts reagent |
| CN109652407A (en) * | 2019-01-31 | 2019-04-19 | 华南理工大学 | A kind of extracting method of microalgae cell total serum IgE |
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Application publication date: 20111123 |