CN102344921A - Method for purifying genomic deoxyribonucleic acid (DNA) of plant tissues by using goldmag particles - Google Patents
Method for purifying genomic deoxyribonucleic acid (DNA) of plant tissues by using goldmag particles Download PDFInfo
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Abstract
The invention aims to provide a method for simply and quickly purifying genomic deoxyribonucleic acid (DNA) of plant tissues by using a magnetic nano composite material. The method comprises the following steps of: performing sample lysis by using two kinds of lysis solution, wherein the first lysis solution comprises PVP (poly vinyl pyrrolidone) with mass-to-volume ratio of 2 to 5 percent, 2 to 5 percent of CTAB (cetyl trimethyl ammonium bromide), and beta-mercaptoethanol with volume fraction of 0.5 to 2 percent, and the second lysis solution comprises 3 to 6M of guanidine hydrochloride and Triton X-100 with volume fraction of 2 to 5 percent; blending, and performing full lysis for 5 to 30 min in a water bath; and adding 10 to 50 microliters of 10 mg/ml RNase solution, and thus obtaining the sample lysis solution with the genomic DNA. The purification method is easy to operate, has quick purification process, and has the advantages of high purification rate, high DNA integrity, high purity and the like. According to the method, fractional tissues lysis is not required, and the requirement of extracting the genomic DNA of various plants and different places is met to the great extent.
Description
Technical field
The present invention relates to a kind of method of the plant tissue genomic dna with gold-magnetic particles purifying broad variety plant and different sites.
Background technology
Utilization has the direct transformation receptor of donor DNA of plants of goal gene, and screening purpose character variation offspring is the practical application on agricultural of plant genetic engineering and molecular biotechnology.Extracting the higher genomic dna of quality from the plant of different genera and different sites thereof, to carry out downstream experiment such as subsequent P CR reaction be a key link, and the method for foundation has higher requirements to the purity and the concentration of the genomic dna that extraction obtains.At present, the plant tissue genome DNA extracting method all has very strong specific aim, and mostly can only from fresh plant leaf, extract, can bring significant limitation to downstream application since like this.
The nucleic acid purification method can be divided into two kinds substantially, and a kind of is phenol chloroform extracting liquid phase method of purification, and a kind of is the solid phase method of purification.The phenol chloroform extraction method mainly distributes through organic phase-water removes the purpose that impurity reaches purification of nucleic acid.Solid phase purifying rule combines with the non-special reversibility of solid-phase media through nucleic acid, thereby reaches the purpose of purification of nucleic acid, and its combination comprises that mainly absorption combines and IX combines.
There are some shortcomings in traditional several plant genome DNA extracting method.Need the extracting and centrifugal repeatedly of phenol chloroform, the time of labor, human and material resources like CTAB method and SDS method.In addition, the bibliographical information of existing plant genome DNA purifying all is the fresh blade to certain kind of plant basically, like the method targetedly to exploitations such as the blade of tobacco, tomato leafs.Because the different tissues material of tissue of different plants or same plant, because differences such as its chemical ingredients, weave construction need be selected diverse ways or make some particular processing when extracting genomic dna, this brings great inconvenience for experiment.Therefore, research is general, to be suitable for the method for plant different sites tissue gene group DNA fast also extremely important.
Summary of the invention
The purpose of this invention is to provide a kind of magnetic nanometer composite material that utilizes and carry out method easy, fast purifying plant tissue genomic dna, with solve background technology complicated operation, purifying rate low, do not have a problem such as versatility; Tissue and same plant different tissues applicable to the different plants of purifying.
Technical scheme of the present invention:
A kind of method with gold-magnetic particles purifying plant tissue genomic dna may further comprise the steps:
1) preparation contains the sample dissociation liquid of genomic dna
Get the plant tissue sample; Two kinds of lysates that add 100～800 μ l respectively; Wherein containing mass volume ratio in first kind of lysate is that 2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB) and volume(tric)fraction are 0.5%～2% beta-mercaptoethanol, contains 3～6M Guanidinium hydrochloride in second kind of lysate, volume(tric)fraction is 2%～5%Triton X-100; Behind the mixing in water-bath abundant cracking 5～30min, add the 10mg/ml RNase solution of 10～50 μ l again, obtain containing the sample dissociation liquid of genomic dna; (unit of " mass volume ratio " is g/ml, the quality of expression solute and the ratio of the volume of this kind lysate)
2) with centrifugal after the chloroform extracting
In said sample dissociation liquid, carry out the sample extracting with 0.6ml～1.2ml chloroform, the centrifugal 5min of 8000rpm～13000rpm then obtains containing the supernatant of genomic dna;
3) combine
Get 400～1000 μ g gold-magnetic particles and mix, under the effect of binding buffer liquid, form the solution that contains gold-magnetic particles-genomic dna mixture with the supernatant that contains genomic dna;
4) clean
Solution to containing gold-magnetic particles-genomic dna mixture carries out magnetic resolution, and abandoning supernatant adds the scavenging solution mixing again, and magnetic resolution is abandoned supernatant, obtains gold-magnetic particles-genomic dna mixture;
5) wash-out
With elutriant and gold-magnetic particles-genomic dna mixture mixing, genomic dna is forwarded to the elutriant from gold-magnetic particles, magnetic resolution is clarified until supernatant, collects supernatant, and the gained supernatant is the genomic dna solution that purifying obtains.
Above-mentioned steps 1) containing mass and size concentration in first kind of lysate described in is that 2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB) and volume(tric)fraction are 0.5%～2% beta-mercaptoethanol, contains 3～6M Guanidinium hydrochloride in second kind of lysate, volume(tric)fraction is 2%～5%Triton X-100 and 0.005～0.01M YD 30 (EDTA), 0.5～2M NaCl and 0.05～0.2M Tris-HCl.
Above-mentioned steps 3) the preferred polyoxyethylene glycol of binding buffer liquid in and the mixing solutions of sodium-chlor, wherein the final concentration of sodium-chlor is 1M～3M, and the molecular weight of polyoxyethylene glycol (PEG) is between 6000～10000, and concentration is 10%～30%; Also can select known other binding buffer liquid.
Above-mentioned steps 5) elutriant described in preferably contains TE solution or the nuclease free water of 10mM Tris-HCl and 1mM EDTA, pH=8.0.
Above-mentioned steps 2) adopt chloroform only need carry out a sample extracting.
Above-mentioned steps 4) scavenging solution can adopt 50%～85% ethanol described in.
Above-mentioned steps 1) optimum temps of water-bath described in is 50～65 ℃.
The present invention uses through two kinds of lysates collocation of employing, need not substep cracking tissue, greatly the satisfied extraction to various plants and different sites genomic dna thereof of degree.
1, the present invention has used two kinds of different lysates---CTAB cracking and guanidinesalt cracking; Adopt two kinds of lysate collocation to use cracking plant tissue cell; Can include the plant of most of kind and the plant tissue of different sites; Satisfy the demand of extracting high-quality DNA from the various plants different sites, the DNA purity that obtains is high, does not contain the suppressor factor that suppresses the downstream PCR reaction.
2, do not need extracting repeatedly and centrifugal, the physical property that has reduced in the mechanical use is sheared, and has also avoided the destruction of centrifugal process to genomic dna.
3, do not need the substep cracking, step is easy, and the time is short, and all processes only needs about 35～60min (according to different samples, the cracked asynchronism(-nization)), thereby has shortened the time of extracting DNA greatly.
4, this purification process is easy and simple to handle, and purge process is quick, and the genomic dna quality that obtains is expensive cheap; It is high to have the purifying rate, DNA good in integrity, purity advantages of higher.
5, can be suitable for the extraction of various plants genomic dna, like tobacco, tomato, aloe etc.; Also can be used for the extraction of same plant different tissues genomic dna, like root, stem, blade, tissues such as seedling.
Description of drawings
Fig. 1 is the electrophorogram of the genomic dna that extracts of the present invention;
Swimming lane M wherein: λ Hind III;
Swimming lane 1: the genomic dna that from tobacco leaf, extracts;
Swimming lane 2: the genomic dna that from tomato seedling, extracts
Swimming lane 3: the genomic dna that from the tobacco root, extracts
Swimming lane 4: the genomic dna that from tobacco stem, extracts.
Embodiment
Utilize magnetic carrier to carry out the thinking that nucleic acid purification is a kind of DNA purifying of getting up of development in recent years; Promptly utilize nanometer or micron-size spherical particles to come nonspecific absorption nucleic acid as solid-phase media with superparamagnetism; Under the effect of externally-applied magnetic field, impurity such as DNA-magnetic particle complex body and protein, polysaccharide are separated; Can save repeatedly numerous and diverse operations such as centrifugal, filtration; Reduced the damage that too much mechanical shearing effect causes nucleic acid, and after removing magnetic field, magnetic particle is dispersed in the solution very soon.Because it has characteristics such as easy and simple to handle, purity height, the parent who more and more receives people looks at, and it easily is automated, and has satisfied the demand of operation such as the high-throughput of building the storehouse in the sample on a large scale.
Gold-magnetic particles is meant that the aggregate with magnetic nano-particle or magnetic nano-particle is a nuclear; Magnetic composite particle in precious metal shell formation such as the surperficial parcel of nuclear simple substance gold and silver; Refer to that also the aggregate with magnetic nano-particle or magnetic nano-particle is a nuclear, at the magnetic composite particle of noble metal formation such as nuclear surface-assembled nanometer gold and silver.Said magnetic nano-particle comprises Fe
3O
4, γ-Fe
2O
3Deng the oxide particle of iron, simple substance Fe, Co, Ni particle, or the positive wustite particle that forms by Fe and other metallic element; The aggregate of magnetic nano-particle is meant after modifying the oxide particle aggregate of iron such as the Fe3O4 that forms, γ-Fe2O3; The simple substance Fe, Co, the Ni particle agglomeration that after modifying, form, or the positive wustite particle agglomeration of Fe that after modifying, forms and the formation of other metallic element.
Is that the method for gold-magnetic particles purifying plant tissue genomic dna is further explained below in conjunction with embodiment to the present invention.
Embodiment 1: with the method for gold-magnetic particles purified genomic dna from tobacco leaf
1) plant sample cracking
1.1) get fresh plant leaf tissue 100mg and clean up with zero(ppm) water, surface-moisture is dried, shred and be placed in the 2ml centrifuge tube, add liquid nitrogen grinding., tissue adds 100 μ l lysates 1 [2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB), 0.5%～2% beta-mercaptoethanol, 0.005～0.01M YD 30 (EDTA), 0.5～2M NaCl before thawing; 0.05～0.2M Tris-HCl] and 100 μ l lysates 2 (3～6M GuHCl, 2%～5%Triton X-100), 65 ℃ of water-bath 25min behind the mixing (during every put upside down centrifuge tube 1 time) at a distance from 5min.
1.2) the RNase solution of adding 10 μ l 10mg/ml in centrifuge tube, the pressure-vaccum mixing in 37 ℃ of water-bath 5min, takes out centrifuge tube.
2) chloroform extracting
In centrifuge tube, add the 0.7ml chloroform, composition in the abundant mixing centrifuge tube of jog 5min, in room temperature, the centrifugal 5min of 8000rpm～13000rpm.
3) combine
3.1) fully shake up gold-magnetic particles, pipette 100 μ l gold-magnetic particles in a new 2ml centrifuge tube with pipettor, magnetic resolution 1min abandons supernatant.Take out centrifuge tube from magnetic separator, add 200 μ l and combine liquid, pressure-vaccum mixing.
3.2) from whizzer, take out centrifuge tube, draw supernatant (careful as far as possible absorption of this step gone up honest and upright and thrifty 200 μ l, is not drawn to organic phase) in step 3.1) centrifuge tube in, the pressure-vaccum mixing fully combines in room temperature, room temperature leaves standstill 5min, magnetic resolution 2min abandons supernatant.
4) clean
4.1) in centrifuge tube, add 200 μ l scavenging solutions 1, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.
4.2) in centrifuge tube, add 200 μ l scavenging solutions 2, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.Repeat this step once.
5) wash-out
Open centrifuge tube lid, place on the magnetic separator room temperature to dry 5min centrifuge tube.
In centrifuge tube, add 100 μ l elutriants (10mM Tris-HCl, 1mM EDTA, pH=8.0), pipettor piping and druming mixing is in 60 ℃ of water-bath 5min.Magnetic resolution is clarified until supernatant, and supernatant is the genomic dna of purifying, and supernatant is transferred in another centrifuge tube, can directly be used for subsequent experimental or in-20 ℃ of cryopreservation.
Record the result according to ultraviolet spectrophotometer and show that from the tobacco leaf of 100mg, can obtain the genomic dna of 15～30 μ g, purity is between 1.7～1.9.
Embodiment 2: with the method for gold-magnetic particles purified genomic dna from tomato seedling
1) plant sample cracking
1.1) get the fresh tomato seedling and organize 100mg to clean up with zero(ppm) water, surface-moisture is dried, shred and be placed in the 2ml centrifuge tube, add liquid nitrogen grinding., tissue adds 200 μ l lysates 1 [2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB), 0.5%～2% beta-mercaptoethanol, 0.005～0.01M YD 30 (EDTA), 0.5～2M NaCl before thawing; 0.05～0.2M Tris-HCl] and 200 μ l lysates 2 (3～6M GuHCl, 2%～5%Triton X-100), 65 ℃ of water-bath 25min behind the mixing (during every put upside down centrifuge tube 1 time) at a distance from 5min.
1.2) the RNase solution of adding 10 μ l 10mg/ml in centrifuge tube, the pressure-vaccum mixing in 37 ℃ of water-bath 5min, takes out centrifuge tube.
2) chloroform extracting
In centrifuge tube, add the 0.8ml chloroform, composition in the abundant mixing centrifuge tube of jog 5min, in room temperature, the centrifugal 5min of 8000rpm～13000rpm.
3) combine
3.1) fully shake up gold-magnetic particles, pipette 100 μ l gold-magnetic particles in a new 2ml centrifuge tube with pipettor, magnetic resolution 1min abandons supernatant.Take out centrifuge tube from magnetic separator, add 200 μ l and combine liquid, pressure-vaccum mixing.
3.2) from whizzer, take out centrifuge tube, draw supernatant (careful as far as possible absorption of this step gone up honest and upright and thrifty 200 μ l, is not drawn to organic phase) in step 3.1) centrifuge tube in, the pressure-vaccum mixing fully combines in room temperature, room temperature leaves standstill 5min, magnetic resolution 2min abandons supernatant.
4) clean
4.1) in centrifuge tube, add 200 μ l scavenging solutions 1, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.
4.2) in centrifuge tube, add 200 μ l scavenging solutions 2, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.Repeat this step once.
5) wash-out
Open centrifuge tube lid, place on the magnetic separator room temperature to dry 5min centrifuge tube.
In centrifuge tube, add 100 μ l elutriants (10mM Tris-HCl, 1mM EDTA, pH=8.0), pipettor piping and druming mixing is in 60 ℃ of water-bath 5min.Magnetic resolution is clarified until supernatant, and supernatant is the genomic dna of purifying, and supernatant is transferred in another centrifuge tube, can directly be used for subsequent experimental or in-20 ℃ of cryopreservation.
Record the result according to ultraviolet spectrophotometer and show that from the tomato seedling of 100mg, can obtain the genomic dna of 10～20 μ g, purity is between 1.7～1.9.
Embodiment 3: with the method for gold-magnetic particles purified genomic dna from the tobacco root
1) plant sample cracking
1.1) get fresh tobacco root 100mg and clean up with zero(ppm) water, surface-moisture is dried, shred and be placed in the 2ml centrifuge tube, add liquid nitrogen grinding., tissue adds 400 μ l lysates 1 [2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB), 0.5%～2% beta-mercaptoethanol, 0.005～0.01M YD 30 (EDTA), 0.5～2M NaCl before thawing; 0.05～0.2M Tris-HCl] and 400 μ l lysates 2 (3～6M GuHCl, 2%～5%Triton X-100), 65 ℃ of water-bath 25min behind the mixing (during every put upside down centrifuge tube 1 time) at a distance from 5min.
1.2) the RNase solution of adding 10 μ l 10mg/ml in centrifuge tube, the pressure-vaccum mixing in 37 ℃ of water-bath 5min, takes out centrifuge tube.
2) chloroform extracting
In centrifuge tube, add the 0.6ml chloroform, composition in the abundant mixing centrifuge tube of jog 5min, in room temperature, the centrifugal 5min of 8000rpm～13000rpm.
3) combine
3.1) fully shake up gold-magnetic particles, pipette 100 μ l gold-magnetic particles in a new 2ml centrifuge tube with pipettor, magnetic resolution 1min abandons supernatant.Take out centrifuge tube from magnetic separator, add 200 μ l and combine liquid, pressure-vaccum mixing.
3.2) from whizzer, take out centrifuge tube, draw supernatant (careful as far as possible absorption of this step gone up honest and upright and thrifty 200 μ l, is not drawn to organic phase) in step 3.1) centrifuge tube in, the pressure-vaccum mixing fully combines in room temperature, room temperature leaves standstill 5min, magnetic resolution 2min abandons supernatant.
4) clean
4.1) in centrifuge tube, add 200 μ l scavenging solutions 1, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.
4.2) in centrifuge tube, add 200 μ l scavenging solutions 2, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.Repeat this step once.
5) wash-out
Open centrifuge tube lid, place on the magnetic separator room temperature to dry 5min centrifuge tube.
In centrifuge tube, add 100 μ l elutriants (10mM Tris-HCl, 1mM EDTA, pH=8.0), pipettor piping and druming mixing is in 60 ℃ of water-bath 5min.Magnetic resolution is clarified until supernatant, and supernatant is the genomic dna of purifying, and supernatant is transferred in another centrifuge tube, can directly be used for subsequent experimental or in-20 ℃ of cryopreservation.
Record the result according to ultraviolet spectrophotometer and show, from the root of the tobacco of 100mg, can obtain the genomic dna of 5～10 μ g, purity is between 1.7～1.9.
Embodiment 4: with the method for gold-magnetic particles purified genomic dna from tobacco stem
1) plant sample cracking
1.1) get fresh tobacco stem 100mg and clean up with zero(ppm) water, surface-moisture is dried, shred and be placed in the 2ml centrifuge tube, add liquid nitrogen grinding., tissue adds 400 μ l lysates 1 [2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB), 0.5%～2% beta-mercaptoethanol, 0.005～0.01M YD 30 (EDTA), 0.5～2M NaCl before thawing; 0.05～0.2MTris-HCl] and 400 μ l lysates 2 (3～6M GuHCl, 2%～5%Triton X-100), 65 ℃ of water-bath 25min behind the mixing (during every put upside down centrifuge tube 1 time) at a distance from 5min.
1.2) the RNase solution of adding 10 μ l 10mg/ml in centrifuge tube, the pressure-vaccum mixing in 37 ℃ of water-bath 5min, takes out centrifuge tube.
2) chloroform extracting
In centrifuge tube, add the 0.6ml chloroform, composition in the abundant mixing centrifuge tube of jog 5min, in room temperature, the centrifugal 5min of 8000rpm～13000rpm.
3) combine
3.1) fully shake up gold-magnetic particles, pipette 100 μ l gold-magnetic particles in a new 2ml centrifuge tube with pipettor, magnetic resolution 1min abandons supernatant.Take out centrifuge tube from magnetic separator, add 200 μ l and combine liquid, pressure-vaccum mixing.
3.2) from whizzer, take out centrifuge tube, draw supernatant (careful as far as possible absorption of this step gone up honest and upright and thrifty 200 μ l, is not drawn to organic phase) in step 3.1) centrifuge tube in, the pressure-vaccum mixing fully combines in room temperature, room temperature leaves standstill 5min, magnetic resolution 2min abandons supernatant.
4) clean
4.1) in centrifuge tube, add 200 μ l scavenging solutions 1, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.
4.2) in centrifuge tube, add 200 μ l scavenging solutions 2, pipettor pressure-vaccum mixing, magnetic resolution 2min abandons supernatant as far as possible.Repeat this step once.
5) wash-out
Open centrifuge tube lid, place on the magnetic separator room temperature to dry 5min centrifuge tube.
In centrifuge tube, add 100 μ l elutriants (10mM Tris-HCl, 1mM EDTA, pH=8.0), pipettor piping and druming mixing is in 60 ℃ of water-bath 5min.Magnetic resolution is clarified until supernatant, and supernatant is the genomic dna of purifying, and supernatant is transferred in another centrifuge tube, can directly be used for subsequent experimental or in-20 ℃ of cryopreservation.
Record the result according to ultraviolet spectrophotometer and show, from the stem of the tobacco of 100mg, can obtain the genomic dna of 5～10 μ g, purity is between 1.7～1.9.
Claims (7)
1. method with gold-magnetic particles purifying plant tissue genomic dna is characterized in that this method may further comprise the steps:
1) preparation contains the sample dissociation liquid of genomic dna
Get the plant tissue sample; Two kinds of lysates that add 100～800 μ l successively; Wherein containing mass volume ratio in first kind of lysate is that 2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB) and volume(tric)fraction are 0.5%～2% beta-mercaptoethanol, contains 3～6M Guanidinium hydrochloride in second kind of lysate, volume(tric)fraction is 2%～5%Triton X-100; Behind the mixing in water-bath abundant cracking 5～30min, add the 10mg/ml RNase solution of 10～50 μ l again, obtain containing the sample dissociation liquid of genomic dna;
2) with centrifugal after the chloroform extracting
In said sample dissociation liquid, carry out the sample extracting with 0.6ml～1.2ml chloroform, the centrifugal 5min of 8000rpm～13000rpm then obtains containing the supernatant of genomic dna;
3) combine
Get 400～1000 μ g gold-magnetic particles and mix, under the effect of binding buffer liquid, form the solution that contains gold-magnetic particles-genomic dna mixture with the supernatant that contains genomic dna;
4) clean
Solution to containing gold-magnetic particles-genomic dna mixture carries out magnetic resolution, and abandoning supernatant adds the scavenging solution mixing again, and magnetic resolution is abandoned supernatant, obtains gold-magnetic particles-genomic dna mixture;
5) wash-out
With elutriant and gold-magnetic particles-genomic dna mixture mixing, genomic dna is forwarded to the elutriant from gold-magnetic particles, magnetic resolution is clarified until supernatant, collects supernatant, and the gained supernatant is the genomic dna solution that purifying obtains.
2. the method with gold-magnetic particles purifying plant tissue genomic dna according to claim 1; It is characterized in that: containing mass and size concentration in first kind of lysate described in the step 1) is that 2%～5% Vinylpyrrolidone polymer (PVP), 2%～5% cetyl trimethylammonium bromide (CTAB) and volume(tric)fraction are 0.5%～2% beta-mercaptoethanol, contains 3～6M Guanidinium hydrochloride in second kind of lysate, volume(tric)fraction is 2%～5%Triton X-100 and 0.005～0.01M YD 30 (EDTA), 0.5～2M NaCl and 0.05～0.2M Tris-HCl.
3. method according to claim 2; It is characterized in that: the binding buffer liquid in the step 3) is the mixing solutions of polyoxyethylene glycol and sodium-chlor; Wherein the final concentration of sodium-chlor is 1M～3M, and the molecular weight of polyoxyethylene glycol (PEG) is between 6000～10000, and concentration is 10%～30%.
4. method according to claim 3 is characterized in that: elutriant described in the step 5) is TE solution or the nuclease free water that contains 10mM Tris-HCl and 1mM EDTA, pH=8.0.
5. method according to claim 4 is characterized in that: step 2) adopt chloroform only to carry out a sample extracting.
6. method according to claim 5 is characterized in that: scavenging solution described in the step 4) is 50%～85% ethanol.
7. method according to claim 6 is characterized in that: the temperature of water-bath described in the step 1) is 50～65 ℃.
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| CN102618532A (en) * | 2012-05-02 | 2012-08-01 | 易春 | Kit for extracting genome DNA (Deoxyribose Nucleic Acid) from plant leaves based on paramagnetic particle method and extracting method thereof |
| CN103952400A (en) * | 2014-04-23 | 2014-07-30 | 西安金磁纳米生物技术有限公司 | Method for purifying total nucleic acid in micro tissues of animals by using gold magnetic particles |
| CN104630208A (en) * | 2015-02-11 | 2015-05-20 | 杭州百迈生物技术有限公司 | Kit and method for extracting genome DNA (deoxyribonucleic acid) |
| CN104630208B (en) * | 2015-02-11 | 2018-01-02 | 杭州百迈生物股份有限公司 | Extract the kit and extracting method of genomic DNA |
| CN108795924A (en) * | 2017-12-06 | 2018-11-13 | 宁夏农林科学院 | A kind of quick, simple plant genome DNA extracting method |
| CN110358762A (en) * | 2019-07-26 | 2019-10-22 | 云南省农业科学院热区生态农业研究所 | A kind of kit and its extracting method extracting plant leaf blade genomic DNA |
| EP4163369A1 (en) | 2021-10-08 | 2023-04-12 | AXAGARIUS GmbH & Co. KG | Use of mixtures of polyvinylpyrrolidone and ammonium sulfate in the isolation of nucleic acids |
| CN115058415A (en) * | 2022-08-18 | 2022-09-16 | 南京集思慧远生物科技有限公司 | Rapid, high-quality and universal genome DNA extraction kit and DNA extraction method |
| CN115058415B (en) * | 2022-08-18 | 2022-12-09 | 南京集思慧远生物科技有限公司 | Rapid, high-quality and universal genome DNA extraction kit and DNA extraction method |
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