CN102453709A - Extraction process for plant RNA - Google Patents
Extraction process for plant RNA Download PDFInfo
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- CN102453709A CN102453709A CN2010105105853A CN201010510585A CN102453709A CN 102453709 A CN102453709 A CN 102453709A CN 2010105105853 A CN2010105105853 A CN 2010105105853A CN 201010510585 A CN201010510585 A CN 201010510585A CN 102453709 A CN102453709 A CN 102453709A
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Abstract
The invention relates to an extraction process for plant RNA, belongs to the technical field of chemical engineering, and specifically relates to an extraction process for the plant RNA. With the present invention, the extraction process for the plant RNA is provided, wherein the extraction process has characteristics of simple operation and mild reaction conditions. The process of the present invention adopts the following technical scheme, wherein the process steps comprise that: components of a RNA extraction buffer comprise: 2-4 mol/L of guanidinium isothiocyanate, 25-35 mmol/L of sodium citrate (pH value of 7.0), 0.5-1.0% (W/V) of sodium dodecyl sarcosinate, and 2-4% (V/V) of beta-mercaptoethanol; the pre-cooling extraction buffer is added to a centrifuge tube, and the centrifuge tube is placed on ice; sarcozygium xanthoxylon bunge leaves and sarcozygium xanthoxylon bunge roots are taken, and completely grinded in a mortar filled with excess liquid nitrogen, a pre-cooling medicine spoon is adopted to transfer the powder of the sarcozygium xanthoxylon bunge leaves and the sarcozygium xanthoxylon bunge roots to a centrifuge tube, a vigorous shaking treatment is performed for 10 minutes, and a standing treatment is performed for 5 minutes on the ice; sodium acetate, water-saturated phenol and a mixing solution comprising chloroform and isoamyl alcohol are added, then the mixture is uniformly shaken, and is centrifuged for 20 minutes, wherein a volume ratio of the chloroform to the isoamyl alcohol is 20:1-24:1; 70-80% ethanol is added, the resulting RNA precipitation is washed, and a drying treatment is performed for 10 minutes at a room temperature to obtain the finished product.
Description
Technical field:
The invention belongs to chemical technology field, more particularly, relate to a kind of extraction process of plant RNA.
Background technology:
Because the abundant and complicated component of plant cell wall is rich in secondary metabolite such as tannin, terpenes, pigment and phenol and macromole such as protein and polysaccharide in the cell, these not only influence the RNA extraction efficiency, and disturb thereafter rt and enzyme to cut.In addition, RNA very easily receives the influence of endogenous or exogenous RNA enzyme and degrades and receive the pollution of protein and DNA etc. and reduce the quality of RNA, and it is difficult therefore from plant, to extract RNA.
Be grown in the strong xerophyte overlord in arid desert district, secondary metabolite content is high in its body, yet, present stage, also there is not producer from the overlord, to carry out the extraction of RNA.
Summary of the invention:
The present invention is exactly to the problems referred to above, and a kind of extraction process of simple to operate, plant RNA that reaction conditions is gentle is provided.
In order to realize above-mentioned purpose of the present invention, the present invention adopts following technical scheme, and process step is:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 2～4mol/L, Trisodium Citrate (pH 7.0) 25～35mmol/L, sarcosyl 0.5～1.0% (W/V), beta-mercaptoethanol 2～4% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
Beneficial effect of the present invention:
1. the present invention's weak point simple to operate, consuming time;
2. the RNA of the present invention's extraction receives the degree of pollutions such as polysaccharide, polyphenol and protein little, and purity is high;
3. the RNA of the present invention's extraction is complete, pure, does not have degraded;
4. output of the present invention is 257.6 μ g/g.
Embodiment:
Process step of the present invention is:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 2～4mol/L, Trisodium Citrate (pH 7.0) 25～35mmol/L, sarcosyl 0.5～1.0% (W/V), beta-mercaptoethanol 2～4% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
Embodiment 1:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 4mol/L, Trisodium Citrate (pH 7.0) 35mmol/L, sarcosyl 0.5% (W/V), beta-mercaptoethanol 2% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
Embodiment 2:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 2mol/L, Trisodium Citrate (pH 7.0) 25mmol/L, sarcosyl 1.0% (W/V), beta-mercaptoethanol 4% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
Embodiment 3:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 3mol/L, Trisodium Citrate (pH 7.0) 25mmol/L, sarcosyl 0.75% (W/V), beta-mercaptoethanol 2% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
Claims (4)
1. the extraction process of a plant RNA is characterized in that, the present invention adopts following technical scheme, and process step is:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 2～4mol/L, Trisodium Citrate (pH 7.0) 25～35mmol/L, sarcosyl 0.5～1.0% (W/V), beta-mercaptoethanol 2～4% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
2. the extraction process of a kind of plant RNA according to claim 1 is characterized in that, process step is:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 4mol/L, Trisodium Citrate (pH 7.0) 35mmol/L, sarcosyl 0.5% (W/V), beta-mercaptoethanol 2% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
3. the extraction process of a kind of plant RNA according to claim 1 is characterized in that, process step is:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 2mol/L, Trisodium Citrate (pH 7.0) 25mmol/L, sarcosyl 1.0% (W/V), beta-mercaptoethanol 4% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends; Centrifugal, abandon supernatant, add 70～80% ethanol, washing RNA deposition, dry 10min gets product under the room temperature.
4. the extraction process of a kind of plant RNA according to claim 1 is characterized in that, process step is:
The composition that RNA extracts damping fluid has: guanidinium isothiocyanate 3mol/L, Trisodium Citrate (pH 7.0) 25mmol/L, sarcosyl 0.75% (W/V), beta-mercaptoethanol 2% (V/V);
The extraction damping fluid of getting precooling joins in the centrifuge tube, places on ice; Get overlord's blade, root system and in the mortar of excessive liquid nitrogen is housed, fully grind, with the spoon of precooling overlord's blade, root system powder are changed in the centrifuge tube, concuss 10min leaves standstill 5min on ice; Adding sodium-acetate, water-saturated phenol, volume ratio are 20: 1～24: 1 chloroform and primary isoamyl alcohol mixing solutions, shake up centrifugal 20min; Draw supernatant and change in another centrifuge tube, add the Virahol of precooling, at-10～-20 ℃ of settle 1.5h; Centrifugal, abandon supernatant, be inverted empty doing, add 70～80% ethanol, deposition suspends.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010105105853A CN102453709A (en) | 2010-10-19 | 2010-10-19 | Extraction process for plant RNA |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010105105853A CN102453709A (en) | 2010-10-19 | 2010-10-19 | Extraction process for plant RNA |
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| CN102453709A true CN102453709A (en) | 2012-05-16 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN2010105105853A Pending CN102453709A (en) | 2010-10-19 | 2010-10-19 | Extraction process for plant RNA |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102899318A (en) * | 2012-10-31 | 2013-01-30 | 四川省林业科学研究院 | Method for extracting nanmu RNA (Ribonucleic Acid) |
| CN110760508A (en) * | 2019-05-08 | 2020-02-07 | 漯河市农业科学院 | Method for extracting total RNA of wheat |
| CN114426970A (en) * | 2022-03-17 | 2022-05-03 | 黑龙江省科学院高技术研究院 | Method for extracting RNA (ribonucleic acid) suitable for poplar |
-
2010
- 2010-10-19 CN CN2010105105853A patent/CN102453709A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102899318A (en) * | 2012-10-31 | 2013-01-30 | 四川省林业科学研究院 | Method for extracting nanmu RNA (Ribonucleic Acid) |
| CN110760508A (en) * | 2019-05-08 | 2020-02-07 | 漯河市农业科学院 | Method for extracting total RNA of wheat |
| CN114426970A (en) * | 2022-03-17 | 2022-05-03 | 黑龙江省科学院高技术研究院 | Method for extracting RNA (ribonucleic acid) suitable for poplar |
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Application publication date: 20120516 |