WO2001023348A1 - Stabilized guanidine thiocyanate and the use thereof - Google Patents
Stabilized guanidine thiocyanate and the use thereof Download PDFInfo
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- WO2001023348A1 WO2001023348A1 PCT/EP2000/009342 EP0009342W WO0123348A1 WO 2001023348 A1 WO2001023348 A1 WO 2001023348A1 EP 0009342 W EP0009342 W EP 0009342W WO 0123348 A1 WO0123348 A1 WO 0123348A1
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- guanidine thiocyanate
- stabilized
- aqueous solution
- thiocyanate according
- thiourea
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C279/00—Derivatives of guanidine, i.e. compounds containing the group, the singly-bound nitrogen atoms not being part of nitro or nitroso groups
- C07C279/02—Guanidine; Salts, complexes or addition compounds thereof
Definitions
- the present invention relates to stabilized guanidine thiocyanate and its use as a chaotropic substance in the isolation of nucleic acids from biological material.
- EP-PS 389 063 describes a process for isolating nucleic acids from complex biological starting material, the starting material being mixed with a chaotropic substance and a nucleic acid-binding solid phase (such as SiO ⁇ ), the solid phase Separates nucleic acid complexes and, if necessary, elutes the nucleic acid from the corresponding complexes after washing.
- a chaotropic substance which changes the secondary, tertiary or quaternary structure of the protein or nucleic acid and at least leaves the primary structure intact
- guanidine salts and in particular guanidine thiocyanate are preferably recommended.
- guanidine thiocyanate in a mixture with detergents as a chaotropic substance.
- Guanidine thiocyanate is usually used in the form of an aqueous solution, so-called kits, which usually also contain buffer salts and auxiliary reagents.
- kits which usually also contain buffer salts and auxiliary reagents.
- a disadvantage of the use of guanidine thiocyanate in the isolation of nucleic acids from biological material is the fact that guanidine thiocyanate, particularly in the form of the aqueous solutions, is extremely sensitive to light and temperature.
- the present invention was therefore based on the object of providing a stabilized guanidine thiocyanate which has sufficient light and temperature stability and with which no biochemical interactions with the biological starting materials can be determined at the same time.
- the guanidine thiocyanate contains 5 to 5000 ppm, based on the weight of the guanidine thiocyanate, thiourea and / or N-acetylcysteine.
- the guanidine thiocyanate stabilized according to the invention thus contains 5 to 5,000 ppm, preferably 10 to 100 ppm, (based on the weight of the guanidine thiocyanate) thiourea and N-acetylcysteine.
- the guanidine thiocyanate can be present both in solid form and as an aqueous solution which preferably has a concentration of 30 to 70% by weight and has a pH of 4.5 to 7.0.
- the aqueous solution may also contain the customary buffer salts and auxiliary reagents which are required for isolating nucleic acids from biological materials.
- the aqueous guanidine thiocyanate solution preferably also contains 5 to 50 mmol of buffer salts selected from the group consisting of sodium acetate, sodium phosphate and tris hydrochloride. Dithiothreid (DTT), dithioerythritol (DTE) and mercaptoethanol are preferably used as auxiliary reagents in an amount of 50 to 100 mmol.
- the aqueous guanidine thiocyanate solution also contains 1 to 10% by weight of SiC> 2 particles, which is required as a solid phase in nucleic acid isolation.
- the preparation of the guanidine thiocyanate stabilized according to the invention is relatively unproblematic in that, for example, solid guanidine thiocyanate is mixed with the appropriate amounts of thiourea and / or N-acetylcysteine and then mixed homogeneously.
- aqueous guanidine thiocyanate solutions stabilized guanidine thiocyanate can be converted into the aqueous phase or aqueous guanidine thiocyanate solutions can be mixed with the appropriate amounts of thiourea and / or N-acetylcysteine.
- Another object of the present invention is the use of guanidine thiocyanate stabilized with thiourea and / or N-acetylcysteine as a chaotropic substance in the isolation of nucleic acids from biological material, the guanidine thiocyanate being used in the form of an aqueous solution, if appropriate with the customary additions of buffer salts, auxiliary reagents and solid phase materials.
- the guanidine thiocyanate stabilized according to the invention Due to its excellent light and temperature stability of at least 4 months and the low interaction with the most important biochemical systems, the guanidine thiocyanate stabilized according to the invention has proven itself extremely well in practice.
- a guanidine thiocyanate which has been stabilized with 10 to 20 ppm thiourea and / or N-acetylcysteine has proven particularly suitable.
- sample A3 is mixed with 1 mg of thiourea (approx. 20 ppm) and mixed thoroughly
- sample A4 is mixed with 1 mg of N-acetyl-L-cysteine (approx. 20 ppm) and also mixed thoroughly
- sample A2 , A3 and A4 then stored under the usual daylight at the window. The samples were examined optically for staining.
- sample A5 is stored in the dark, sample A7 mixed with 1 mg thiourea (approx. 10 ppm) and mixed thoroughly, sample A8 mixed with 1 mg N-acetylcysteine (approx. 10 ppm) and also mixed thoroughly, samples A6, A7 and A8 then stored under the usual daylight at the window. The samples were examined optically for staining and, after intensive mixing, for turbidity.
- Example 2 Each 2 g of solutions prepared as in Example 2 are mixed with 100 mg SiO 2 (Aerosil, Akzo). Probe Bl is stored in the dark. Samples B2, B3 and B4 are then stored under the usual daylight at the window. The samples were examined optically for coloring and after intensive mixing after settling of the silica particles for turbidity.
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- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention relates to stabilized guanidine thiocyanate that contains 5 to 5000 ppm, based on the weight of the guanidine thiocyanate, of thiourea and/or N-acetylcysteine. Due to its excellent light and temperature stability and the low interaction with the biochemical systems the stabilized guanidine thiocyanate according to the invention is suitable to be used as a chaotropic substance for the isolation of nucleic acids from biological material.
Description
Stabilisiertes Guanidinthiocyanat und dessen Verwendung Stabilized guanidine thiocyanate and its use
Die vorliegende Erfindung betrifft stabilisiertes Guanidinthiocyanat und dessen Verwendung als chaotrope Substanz bei der Isolierung von Nucleinsäuren aus biologischem Material.The present invention relates to stabilized guanidine thiocyanate and its use as a chaotropic substance in the isolation of nucleic acids from biological material.
Aufgrund der steigenden Bedeutung von DNA- oder RNA-Analysen im medizinischen und biotechnischen Bereich ist die Nachfrage nach einfachen und effektiven Methoden zur Isolierung von Nucleinsäuren aus biologischem Material sehr stark angestiegen.Due to the increasing importance of DNA or RNA analysis in the medical and biotechnical field, the demand for simple and effective methods for isolating nucleic acids from biological material has increased significantly.
So wird bspw. in der EP-PS 389 063 ein Verfahren zur Isolierung von Nucleinsäuren aus komplexem biologischem Ausgangsmaterial beschrieben, wobei man das Ausgangsmaterial mit einer chaotropen Substanz und einer Nucleinsäure bindenden Festphase (wie z. B. SiO^) mischt, die Festphase- Nucleinsäure-Komplexe abtrennt und ggf. nach dem Waschen die Nucleinsäure von den entsprechenden Komplexen eluiert . Als chaotrope Substanz, welche die Sekundär-, Tertiär- oder Quarternärstruktur des Proteins oder der Nucleinsäure ändert und zumindest die Primärstruktur intakt läßt, werden vorzugsweise Guanidinsalze und insbesondere Guanidinthiocyanat empfohlen.For example, EP-PS 389 063 describes a process for isolating nucleic acids from complex biological starting material, the starting material being mixed with a chaotropic substance and a nucleic acid-binding solid phase (such as SiO ^), the solid phase Separates nucleic acid complexes and, if necessary, elutes the nucleic acid from the corresponding complexes after washing. As a chaotropic substance which changes the secondary, tertiary or quaternary structure of the protein or nucleic acid and at least leaves the primary structure intact, guanidine salts and in particular guanidine thiocyanate are preferably recommended.
Auch entsprechend der Veröffentlichung von Piotr Chomczynski et al. in Biotechnics, Vol. 22, Nr. 3 (1997) wird Guanidinthiocyanat im Gemisch mit Detergenzien als chaotrope Substanz beschrieben. Guanidinthiocyanat wird üblicherweise hierbei im Form einer wäßrigen Lösung, sog. Kits, eingesetzt, die üblicherweise noch Puffersalze und Hilfsreagenzien enthalten .
Nachteilig bei der Verwendung von Guanidinthiocyanat bei der Isolierung von Nucleinsäuren aus biologischem Material ist die Tatsache, daß Guanidinthiocyanat, insbesondere in Form der wäßrigen Lösungen, äußerst licht- und temperaturempfindlich ist. Vor allem im schwach sauren Medium (pH 4,5 bis 7,0) kommt es zu unerwünschten Redoxreaktionen, wobei elementarer Schwefel als Zersetzungsprodukt entsteht . Durch die Zersetzungsreaktion, die sich durch eine intensive Gelbfärbung bemerkbar macht, wird das eingesetzte Material in kürzester Zeit unbrauchbar.According to the publication by Piotr Chomczynski et al. Biotechnics, Vol. 22, No. 3 (1997) describes guanidine thiocyanate in a mixture with detergents as a chaotropic substance. Guanidine thiocyanate is usually used in the form of an aqueous solution, so-called kits, which usually also contain buffer salts and auxiliary reagents. A disadvantage of the use of guanidine thiocyanate in the isolation of nucleic acids from biological material is the fact that guanidine thiocyanate, particularly in the form of the aqueous solutions, is extremely sensitive to light and temperature. Undesirable redox reactions occur especially in the weakly acidic medium (pH 4.5 to 7.0), whereby elemental sulfur is formed as a decomposition product. Due to the decomposition reaction, which is noticeable by an intense yellow color, the material used becomes unusable in a very short time.
Zur Lösung dieses Problems wurde bereits vorgeschlagen, den Guanidinthiocyanat -Lösungen Stabilisatoren in Form von Komplexbildnern, wie z. B. EDTA, zuzusetzen. Es hat sich jedoch gezeigt, daß derartige Komplexbildner mit dem biologischen Material unerwünschte biochemische Interaktionen eingehen.To solve this problem, it has already been proposed to stabilize the guanidine thiocyanate solutions in the form of complexing agents, such as. B. EDTA to add. However, it has been shown that such complexing agents enter into undesirable biochemical interactions with the biological material.
Der vorliegenden Erfindung lag daher die Aufgabe zugrunde, ein stabilisiertes Guanidinthiocyanat bereitzustellen, welches eine ausreichende Licht- und Temperaturstabilität aufweist und mit dem gleichzeitig keinerlei biochemische Wechselwirkungen mit den biologischen Ausgangsmaterialien festzustellen ist.The present invention was therefore based on the object of providing a stabilized guanidine thiocyanate which has sufficient light and temperature stability and with which no biochemical interactions with the biological starting materials can be determined at the same time.
Diese Aufgabe wurde erfindungsgemäß dadurch gelöst, daß das Guanidinthiocyanat 5 bis 5 000 ppm bezogen auf das Gewicht des Guanidinthiocyanats , Thioharnstoff und/oder N- Acetylcystein enthält.This object has been achieved according to the invention in that the guanidine thiocyanate contains 5 to 5000 ppm, based on the weight of the guanidine thiocyanate, thiourea and / or N-acetylcysteine.
Es hat sich hierbei überraschenderweise gezeigt, daß durch diesen Zusatz die Licht- und Temperaturstabilität von Guanidinthiocyanat erheblich verbessert wird, ohne dessen Produkteigenschaften, insbesondere die spektroskopischen Eigenschaften, negativ zu beeinflussen.
Das erfindungsgemäß stabilisierte Guanidinthiocyanat enthält also 5 bis 5 000 ppm, vorzugsweise 10 bis 100 ppm, (bezogen auf das Gewicht des Guanidinthiocyanats) Thioharnstoff und N- Acetylcystein . Das Guanidinthiocyanat kann hierbei sowohl in fester Form als auch als wäßrige Lösung vorliegen, die vorzugsweise eine Konzentration von 30 bis 70 Gew.-% aufweist und einen pH-Wert von 4,5 bis 7,0 besitzt.Surprisingly, it has been found that this addition significantly improves the light and temperature stability of guanidine thiocyanate without adversely affecting its product properties, in particular the spectroscopic properties. The guanidine thiocyanate stabilized according to the invention thus contains 5 to 5,000 ppm, preferably 10 to 100 ppm, (based on the weight of the guanidine thiocyanate) thiourea and N-acetylcysteine. The guanidine thiocyanate can be present both in solid form and as an aqueous solution which preferably has a concentration of 30 to 70% by weight and has a pH of 4.5 to 7.0.
Es ist im Rahmen der vorliegenden Erfindung ohne weiteres möglich, daß die wäßrige Lösung noch die üblichen Puffersalze und Hilfsreagenzien enthält, die bei der Nucleinsäuren- Isolierung aus biologischen Materialien erforderlich sind. Vorzugsweise enthält die wäßrige Guanidinthiocyanat -Lösung noch 5 bis 50 mmol Puffersalze ausgewählt aus der Gruppe Natriumacetat , Natriumphosphat und Tris-Hydrochlorid. Als Hilfsreagenzien werden vorzugsweise Dithiothreid (DTT) , Dithioerythrit (DTE) sowie Mercaptoethanol in einer Menge von 50 bis 100 mmol eingesetzt. Gemäß einer bevorzugten Ausführungsform enthält die wäßrige Guanidinthiocyanat-Lösung noch 1 bis 10 Gew.-% an SiC>2-Partikeln, die bei der Nucleinsäuren-Isolierung als Festphase benötigt wird.In the context of the present invention, it is readily possible for the aqueous solution to also contain the customary buffer salts and auxiliary reagents which are required for isolating nucleic acids from biological materials. The aqueous guanidine thiocyanate solution preferably also contains 5 to 50 mmol of buffer salts selected from the group consisting of sodium acetate, sodium phosphate and tris hydrochloride. Dithiothreid (DTT), dithioerythritol (DTE) and mercaptoethanol are preferably used as auxiliary reagents in an amount of 50 to 100 mmol. According to a preferred embodiment, the aqueous guanidine thiocyanate solution also contains 1 to 10% by weight of SiC> 2 particles, which is required as a solid phase in nucleic acid isolation.
Die Herstellung des erfindungsgemäß stabilisierten Guanidinthiocyanats ist relativ unproblematisch, indem bspw. festes Guanidinthiocyanat mit den entsprechenden Mengen an Thioharnstoff und/oder N-Acetylcystein versetzt und anschließend homogen vermischt wird. Zur Herstellung von wäßrigen Guanidinthiocyanat-Lösungen kann bereits stabilisiertes Guanidinthiocyanat in die wäßrige Phase übergeführt werden oder wäßrige Guanidinthiocyanat -Lösungen mit den entsprechenden Mengen an Thioharnstoff und/oder N- Acetylcystein versetzt werden.The preparation of the guanidine thiocyanate stabilized according to the invention is relatively unproblematic in that, for example, solid guanidine thiocyanate is mixed with the appropriate amounts of thiourea and / or N-acetylcysteine and then mixed homogeneously. To prepare aqueous guanidine thiocyanate solutions, stabilized guanidine thiocyanate can be converted into the aqueous phase or aqueous guanidine thiocyanate solutions can be mixed with the appropriate amounts of thiourea and / or N-acetylcysteine.
Ein weiterer Gegenstand der vorliegenden Erfindung ist die Verwendung des mit Thioharnstoff und/oder N-Acetylcystein stabilisierten Guanidinthiocyanats als chaotrope Substanz bei der Isolierung von Nucleinsäuren aus biologischem Material,
wobei das Guanidinthiocyanat in Form einer wäßrigen Lösung ggf. mit den üblichen Zusätzen an Puffersalzen, Hilfsreagenzien und Festphasenmaterialien eingesetzt wird.Another object of the present invention is the use of guanidine thiocyanate stabilized with thiourea and / or N-acetylcysteine as a chaotropic substance in the isolation of nucleic acids from biological material, the guanidine thiocyanate being used in the form of an aqueous solution, if appropriate with the customary additions of buffer salts, auxiliary reagents and solid phase materials.
Aufgrund ihrer ausgezeichneten Licht- und Temperaturstabilität von mindestens 4 Monaten und der geringen Wechselwirkung mit den wichtigsten biochemischen Systemen hat sich das erfindungsgemäß stabilisierte Guanidinthiocyanat hervorragend in der Praxis bewährt .Due to its excellent light and temperature stability of at least 4 months and the low interaction with the most important biochemical systems, the guanidine thiocyanate stabilized according to the invention has proven itself extremely well in practice.
Als besonders gut geeignet hat sich ein Guanidinthiocyanat erwiesen, welches mit 10 bis 20 ppm Thioharnstoff und/oder N- Acetylcystein stabilisiert wurde.A guanidine thiocyanate which has been stabilized with 10 to 20 ppm thiourea and / or N-acetylcysteine has proven particularly suitable.
Die nachfolgenden Beispiele sollen die Erfindung näher veranschaulichen .The following examples are intended to illustrate the invention in more detail.
BeispieleExamples
Beispiel 1example 1
Je 50 g festes Guanidinthiocyanat (Reinheit 99,6 %) wird in eine Laborglasflasche (Schott, Duran, hell) gefüllt. Probe AI wird unter Lichtausschluß gelagert, Probe A3 mit 1 mg Thioharnstoff (ca. 20 ppm) versetzt und intensiv durchmischt, Probe A4 mit 1 mg N-Acetyl-L-Cystein (ca. 20 ppm) versetzt und ebenfalls intensiv durchmischt, Proben A2 , A3 und A4 sodann unter üblicher Tageslichteinstrahlung am Fenster gelagert . Die Proben wurden optisch auf Färbungen untersucht .
50 g solid guanidine thiocyanate (purity 99.6%) is filled into a laboratory glass bottle (Schott, Duran, light). Sample AI is stored in the dark, sample A3 is mixed with 1 mg of thiourea (approx. 20 ppm) and mixed thoroughly, sample A4 is mixed with 1 mg of N-acetyl-L-cysteine (approx. 20 ppm) and also mixed thoroughly, sample A2 , A3 and A4 then stored under the usual daylight at the window. The samples were examined optically for staining.
V Vergleich , E = ErfindungV comparison, E = invention
Beispiel 2Example 2
Je 50 g festes Guanidinthiocyanat (Reinheit 99,6 %) wird in eine Laborglasflasche (Schott, Duran, hell) gefüllt und in 50 g vollentsalztem Wasser (Millipore Laborreinst- wassererzeugungsanlage) gelöst. Probe A5 wird unter Lichtausschluß gelagert, Probe A7 mit 1 mg Thioharnstoff (ca. 10 ppm) versetzt und intensiv durchmischt, Probe A8 mit 1 mg N-Acetylcystein (ca. 10 ppm) versetzt und ebenfalls intensiv durchmischt, Proben A6 , A7 und A8 sodann unter üblicher Tageslichteinstrahlung am Fenster gelagert. Die Proben wurden optisch auf Färbungen und nach intensivem Durchmischen auf Trübungen untersucht .50 g of solid guanidine thiocyanate (purity 99.6%) is poured into a laboratory glass bottle (Schott, Duran, light) and dissolved in 50 g of deionized water (Millipore laboratory ultrapure water production system). Sample A5 is stored in the dark, sample A7 mixed with 1 mg thiourea (approx. 10 ppm) and mixed thoroughly, sample A8 mixed with 1 mg N-acetylcysteine (approx. 10 ppm) and also mixed thoroughly, samples A6, A7 and A8 then stored under the usual daylight at the window. The samples were examined optically for staining and, after intensive mixing, for turbidity.
V Vergleich Erfindung
Beispiel 3V comparison invention Example 3
Je 2 g Lösungen wie unter Beispiel 2 hergestellt werden mit 100 mg Siθ2 (Aerosil, Akzo) versetzt. Probe Bl wird unter Lichtausschluß gelagert. Proben B2 , B3 und B4 werden sodann unter üblicher Tageslichteinstrahlung am Fenster gelagert. Die Proben wurden optisch auf Färbungen und nach intensivem Durchmischen nach Absetzen der Silica Partikel auf Trübungen untersucht .Each 2 g of solutions prepared as in Example 2 are mixed with 100 mg SiO 2 (Aerosil, Akzo). Probe Bl is stored in the dark. Samples B2, B3 and B4 are then stored under the usual daylight at the window. The samples were examined optically for coloring and after intensive mixing after settling of the silica particles for turbidity.
V = Vergleich , E = Erfindung
V = comparison, E = invention
Claims
1. Stabilisiertes Guanidinthiocyanat, dadurch gekennzeichnet, daß es 5 bis 5 000 ppm, bezogen auf das Gewicht des Guanidinthiocyanats, Thioharnstoff und/oder N-Acetylcystein enthält.1. Stabilized guanidine thiocyanate, characterized in that it contains 5 to 5,000 ppm, based on the weight of the guanidine thiocyanate, thiourea and / or N-acetylcysteine.
2. Guanidinthiocyanat nach Anspruch 1, dadurch gekennzeichnet, daß der Gehalt an Thioharnstoff und/oder N-Acetylcystein 10 bis 100 ppm beträgt.2. guanidine thiocyanate according to claim 1, characterized in that the content of thiourea and / or N-acetylcysteine is 10 to 100 ppm.
3. Guanidinthiocyanat nach den Ansprüchen 1 und 2 , dadurch gekennzeichnet, daß es in fester Form vorliegt.3. guanidine thiocyanate according to claims 1 and 2, characterized in that it is in solid form.
4. Guanidinthiocyanat nach den Ansprüchen 1 und 2, dadurch gekennzeichnet, daß es in Form einer wäßrigen Lösung mit einer Konzentration von 30 bis 70 Gew.-% vorliegt.4. guanidine thiocyanate according to claims 1 and 2, characterized in that it is in the form of an aqueous solution with a concentration of 30 to 70 wt .-%.
5. Guanidinthiocyanat nach Anspruch 4 , dadurch gekennzeichnet, daß die wäßrige Lösung einen pH-Wert von 4,5 bis 7,0 aufweist.5. guanidine thiocyanate according to claim 4, characterized in that the aqueous solution has a pH of 4.5 to 7.0.
6. Guanidinthiocyanat nach den Ansprüchen 4 und 5, dadurch gekennzeichnet, daß die wäßrige Lösung noch 5 bis 50 mmol Puffersalze, ausgewählt aus der Gruppe Natriumacetat , Natriumphosphat und Tris-Hydrochlorid, enthält.6. guanidine thiocyanate according to claims 4 and 5, characterized in that the aqueous solution still contains 5 to 50 mmol of buffer salts, selected from the group sodium acetate, sodium phosphate and tris hydrochloride.
7. Guanidinthiocyanat nach den Ansprüchen 4 bis 6, dadurch gekennzeichnet, daß die wäßrige Lösung noch mit 50 bis 100 mmol an Hilfsreagenzien, wie z. B. Dithiothreid (DTT) , Dithioerythrit (DTE) sowie Mercaptoethanol , versetzt ist.7. guanidine thiocyanate according to claims 4 to 6, characterized in that the aqueous solution with 50 to 100 mmol of auxiliary reagents, such as. B. dithiothreid (DTT), dithioerythritol (DTE) and mercaptoethanol.
8. Guanidinthiocyanat nach den Ansprüchen 4 bis 7, dadurch gekennzeichnet, daß die wäßrige Lösung noch 1 bis 108. guanidine thiocyanate according to claims 4 to 7, characterized in that the aqueous solution still 1 to 10
Gew.-% an Siθ2"Partikeln enthält. Verwendung des stabilisierten Guanidinthiocyanats nach den Ansprüchen 1 bis 8 als chaotrope Substanz bei der Isolierung von Nucleinsäuren aus biologischem Material. % By weight of SiO 2 "particles. Use of the stabilized guanidine thiocyanate according to claims 1 to 8 as a chaotropic substance in the isolation of nucleic acids from biological material.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE1999146597 DE19946597A1 (en) | 1999-09-29 | 1999-09-29 | Stabilized guanidine thiocyanate and its use |
| DE19946597.5 | 1999-09-29 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| WO2001023348A1 true WO2001023348A1 (en) | 2001-04-05 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| PCT/EP2000/009342 WO2001023348A1 (en) | 1999-09-29 | 2000-09-25 | Stabilized guanidine thiocyanate and the use thereof |
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| Country | Link |
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| DE (1) | DE19946597A1 (en) |
| WO (1) | WO2001023348A1 (en) |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0389063A2 (en) * | 1989-03-23 | 1990-09-26 | Akzo Nobel N.V. | Process for isolating nucleic acid |
| DE19755960C1 (en) * | 1997-12-16 | 1998-11-26 | Hoechst Ag | Braking open a structure of e.g. microorganism or cell |
-
1999
- 1999-09-29 DE DE1999146597 patent/DE19946597A1/en not_active Withdrawn
-
2000
- 2000-09-25 WO PCT/EP2000/009342 patent/WO2001023348A1/en active Application Filing
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0389063A2 (en) * | 1989-03-23 | 1990-09-26 | Akzo Nobel N.V. | Process for isolating nucleic acid |
| DE19755960C1 (en) * | 1997-12-16 | 1998-11-26 | Hoechst Ag | Braking open a structure of e.g. microorganism or cell |
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| Publication number | Publication date |
|---|---|
| DE19946597A1 (en) | 2001-04-12 |
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