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Stable lysis buffer mixture for extracting nucleic acids

a lysis buffer and buffer mixture technology, applied in the preparation of sugar derivatives, enzymes, sugar derivatives, etc., can solve the problems of troublesome solution stability of corresponding nucleic acids, and achieve the effect of stable use, favorable price and stable concentration

Inactive Publication Date: 2011-04-21
BENDZKO PETER +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0034]The task underlying the invention is thus to provide an improved nucleic acid extraction system which is favourably priced, stable and simple to use, at the same time fulfilling the requirements of a modern nucleic acid extraction system by, inter alia, containing extraction controls.

Problems solved by technology

However, keeping corresponding nucleic acids in suitable concentrations stable in solution is problematic.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

embodiment 1

[0086]Production of a Mixture According to the Invention Described Here for the Lysis of Diagnostic Samples Containing Bacteria

[0087]A solution I of 1.5 M ammonium chloride, 10 mM Tris pH 8, 2% CTAB, 0.5 mg proteinase K / ml and 0.5 mg lysozyme / ml is produced.

[0088]A solution II of 600 μg / ml polyadenyl RNA, 107 copies / ml control DNA fragment plasmid pCONT, 1% trehalose and 50 mM TrisCl, pH 8, is produced.

[0089]Solutions I and II are mixed in a ratio of 40:1.

[0090]The solution is frozen in 400 μl portions in closable 2 ml reaction vessels. Then, these vessels are freeze-dried with the contents and are referred to as extraction tubes bacteria in the further text.

embodiment 2

[0091]Production of a Mixture According to the Invention Described Here for the Lysis of Diagnostic Samples Containing Viruses

[0092]A solution I of 1.5 M ammonium chloride, 10 mM Tris pH 8, 2% CTAB, 0.5 mg proteinase K / ml and 0.5 mg lysozyme / ml is produced.

[0093]A solution II of 600 μg / ml polyadenyl RNA, 107 copies / ml control DNA fragment plasmid pCONT, 108 copies / ml control RNA fragment, 1% trehalose and 50 mM TrisCl, pH 8, is produced.

[0094]Solutions I and II are mixed in a ratio of 40:1.

[0095]The solution is frozen in 400 μl portions in closable 2 ml reaction vessels. Then, these vessels are freeze-dried with the contents and are referred to as extraction tubes virus in the further text.

embodiment 3

[0096]Standardised Extraction of Viral RNA (Influenza A Virus) and Viral DNA (HBV) by Means of the Produced Lysis Mixture via Spin Filters

[0097]Serum samples (200 μl) with the corresponding viruses in a known number of copies (HBV 500 copies per preparation) or with an estimated titre quantity (Influenza A Virus) are used for extraction.

[0098]200 μl serum and 200 μl water are poured into an extraction tube virus with the mixture for virus lysis and there is thorough mixing. There is then incubation for 15 min at 65° C. in an Eppendorf thermo-mixer under continuous shaking, then incubation for 10 min at 95° C. in the Eppendorf thermo-mixer. 400 μl of isopropyl alcohol is added, followed by mixing by means of repeated pipetting up and down The lysate is placed on a spin filter of the firm of Invitek and incubated for one minute at room temperature. After this, it is centrifuged for one minute at 10,000 rpm in an Eppendorf table-top centrifuge. The spin filter is washed twice with a wa...

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Abstract

The invention relates to a lysis buffer mixture that is stable in storage for isolating nucleic acids from biological, preferably diagnostic samples. The mixture is preferably associated with an extraction control. The aim of the invention is to provide an improved nucleic acid extraction system, which is cost-effective, stable and easy to use, thus fulfilling the requirements of a modern nucleic acid extraction system and containing, among other things, extraction controls. The invention relates to a lysis buffer mixture for isolating nucleic acids, said mixture containing non chaotropic salts, a special selection of detergents, a defined quantity of at least one nucleic acid as an extraction control, optionally lytic enzymes, optionally carrier nucleic acids and optionally other additives.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application is the United States National Phase under 35 U.S.C. §371 of PCT International Patent Application No. PCT / DE2009 / 000549, filed on Apr. 20, 2009, and claiming priority to German Patent Application No. 10 2008 020 258.4, filed on Apr. 22, 2008. Those applications are incorporated by reference herein.BACKGROUND OF THE INVENTION1. Field of the Invention[0002]The invention relates to a storage-stable lysis buffer mixture for extraction of nucleic acids from biological, preferably diagnostic samples. It is preferably connected with an extraction control.[0003]Fields of application are molecular biology diagnostics, research, medical practice, gene-based analysis of biotechnological, agricultural and foodstuff products as well as criminal science.[0004]2. Background of the Related Art[0005]A large number of customary lysis buffers for extraction of nucleic acids contain chaotropic ion mixtures as salts. The pertinent methods are ...

Claims

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Application Information

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IPC IPC(8): C07H1/06C12N1/06C12N7/00C12N9/50
CPCC12N15/1003C12N15/1013C12N9/96C12N9/2462C12N9/6424C12N15/1017
Inventor BENDZKO, PETERJOOS, HANS
Owner BENDZKO PETER
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