RU1770358C - Method for plant nucleic acids separation by electrophoresis - Google Patents

Abstract

Usage: biochemistry and molecular biology. Essence: in order to control the speed of movement of nucleic acid fragments in the electrophoretic system, the drug is treated with a silicon compound, for example, sodium metasilicate at a concentration of M, before being added to the gel. This results in a 20% decrease in the speed of movement of nucleic acid fragments in the electromagnetic field compared to control. 1 tab.

Description

The invention relates to the field of plant biochemistry and molecular biology and can be used in the assessment of starting material in rice selection; in solving genetic engineering problems: in the laboratory practice of molecular biological research.

A known method of electrophoresis of biological macromolecules on paper, in starch gels (1), in polyacrylamide gels (2).

These methods are of little use for electrophoresis of nucleic acids (NKs), since they do not allow efficient separation of the macromolecules of the latter; if used, it is difficult to isolate purified nucleic acid fragments from the gel itself.

Methods for electrophoretic separation of nucleic acids using polyacrylamide-agarose, agar gels (3) as well as agarose gels (4) are also known.

The disadvantages of the above methods are:

- the inability to directionally control the speed of movement of nucleic acid fragments in the electrophoretic system, which is caused by the low resolution of the method;

-additional costs of time and money associated with repeated and subsequent operations to separate nucleic acid fragments that are close in molecular characteristics.

Closest to the proposed is a method for electrophoretic separation of nucleic acids (5). which consists in the fact that the studied NA preparation is introduced into the well of a horizontal underwater (flooded with Tris-borate buffer pH 8.0) agarized gel (agarose concentration 0.9%, gel thickness 2 mm, length 20 cm), followed by electrophoresis the reticulum system with a power source is exposed for 5 hours

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electromagnetic field with parameters: current strength - 194 mA, voltage - 82 mV. Then the process is stopped and the results are studied on a transilluminator.

The disadvantages of the method are:

-low resolution with respect to fragments close in molecular weight;

- the impossibility of directional regulation of fragments of NK in an electromagnetic field;

-additional costs of time and money associated with repeated and subsequent operations to separate fragments of nanocrystals close in molecular characteristics.

The aim of the invention is to increase the resolution of the method by reducing the speed of movement of nucleic acid fragments in an electromagnetic field.

This goal is achieved by the fact that in the known method of electrophoretic separation of nucleic acids, which consists in placing the NK preparation in an agarose gel flooded with a buffer and exposing the said preparation to an electromagnetic field, the drug is treated with sodium metasilicate at a concentration of 10-10 before it is introduced into the gel

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The proposed method is carried out as follows.

The studied preparation of deoxyribonucleic acid (DNA) from 12-day-old rice seedlings of the Krasnodar 424 variety was kept in solution, 10 M, M, M, 10 M sodium metasilicate, depending on the experimental version, for 24 hours, then dissolved in 8% a solution of sucrose containing 0.1% bromophenol blue was added to the well of a horizontal underwater agarose gel. Gel

flooded with a layer of Tris-borate buffer, pH 8.0. The agarose concentration in the gel is 0.9%, the gel thickness is 2 mm, and the length is 20 cm. Then, the electromagnetic field was applied to the electrophoretic system using a power source for 5 hours with the following parameters: current strength 194 mA, voltage 82 mV. The progress of electrophoresis was monitored by the movement of bromphenol blue. At the end of the process, the gel was stained in ethidium bromide solution and viewed on a transilluminator. The electrolysis results are presented in the table. As follows from the data presented, treatment of NK with a 10, 10 M sodium metasilicate solution leads to a change in electrophoretic parameters: a decrease in the distance traveled and a decrease in the speed of motion of NK fragments. This, in turn, allows one to separate NC fragments that are close in electrophoretic properties (mass, charge, etc.); isolate from the aggregate of fragments one that is necessary for biochemical and genetically engineered purposes.

Claims (1)

The claims A method for electrophoretic separation of plant nucleic acids, comprising introducing a nucleic acid preparation into an agarose gel and exposing the preparation to an electromagnetic field, characterized in that, in order to increase the resolution of the method by reducing the speed of movement of the nucleic acid fragments in the electromagnetic field, before the preparation is introduced into the gel nucleic acid is treated with sodium metasilicate in a concentration of - M.