Chemistry: molecular biology and microbiology – Apparatus – Including measuring or testing
Reexamination Certificate
2000-03-02
2001-12-04
Redding, David A. (Department: 1744)
Chemistry: molecular biology and microbiology
Apparatus
Including measuring or testing
C435S288100, C435S288600, C210S198200, C210S502100, C210S510100, C210S263000, C210S321750, C210S321840, C095S088000, C096S107000
Reexamination Certificate
active
06326189
ABSTRACT:
BACKGROUND OF THE INVENTION
The present invention relates to a nucleic acid separating vessel for separating the nucleic acid from a liquid sample, to a method of manufacturing the same, and to a method for separating the nucleic acid.
A conventional method for separating and purifying a nucleic acid comprising the steps of isolating the nucleic acid into a liquid sample by making a biological sample and the like soluble in the solution, extracting the nucleic acid with an organic solvent and the like, precipitating the nucleic acid with alcohol and the like, and recovering the nucleic acid, has been used widely.
However, the above conventional method is inadequate for automatic operation, in view of requiring a large amount of labor and learning the operating procedures, and of frequent use of centrifugal separating operation, because it requires a large number of operating steps.
In order to overcome the above problems, a method utilizing an adsorption characteristics of the nucleic acid to a solid phase body has been disclosed. For instance, a description that the nucleic acid combines with glass under the presence of chaotropic salt is disclosed in the Proceedings of National Academy of Science of the USA vol. 76, 615-619 (1979).
Utilizing the characteristics of the nucleic acid to combine with a silicon oxide (silica) under a certain condition, a nucleic acid extracting kit and a nucleic acid automatic extracting apparatus using magnetic silica particles, silica particles, silica fiber or filter, spin column or micro plate containing the above articles, and the like are obtainable commercially.
However, in accordance with the above prior art, a magnet is necessary for separating the magnetic particles from the liquid sample (B/F separation) when the magnetic particles are used, and a centrifugal separating operation is necessary when the glass particles or spin column is used. Accordingly, the operation and composition of the apparatus become composite. In particular, when the magnetic particles were used in a micro plate type, magnetic particle separating mechanisms as same number as the number of wells became necessary, and there were problems such that the composition of the apparatus becomes composite and expensive.
In a case when silica fiber or filter, and micro plate containing them are used, the B/F separation is performed by a filtering operation or a centrifugal operation. When particle size was decreased, or fiber density was increased in order to increase combining ability of these materials with nucleic acid, there were problems such that passing velocity of the liquid sample was decreased, and the velocity of the B/F separation was decreased.
SUMMARY OF THE INVENTION
One of the objects of the present invention is to realize a nucleic acid separating vessel, which is capable of suppressing decrease of the B/F separating velocity with maintaining the nucleic acid combining ability high.
Other one of the objects of the present invention is to realize a method of manufacturing the nucleic acid separating vessel, which is capable of suppressing decrease of the B/F separating velocity with maintaining the nucleic acid combining ability high.
Other one of the objects of the present invention is to realize a method for separating nucleic acid using the nucleic acid separating vessel, which is capable of suppressing decrease of the B/F separating velocity with maintaining the nucleic acid combining ability high.
In order to realize these objects, the present invention is composed as follows:
(1) A nucleic acid separating vessel for separating the nucleic acid from a biological sample provided with a silicon oxide composite particle structural body as a separating means for separating the nucleic acid from a biological sample; wherein a silicon oxide composite particle is formed by making silicon oxide group particles form a composite with surface of a particle, which is larger than the silicon oxide group particle and becomes a nucleus; and the plural silicon oxide composite particles are combined each other three dimensionally to form the silicon oxide composite particle structural body.
(2) Preferably, the plural nucleic acid separating vessels described in the above (1) are arranged on a same supporting plate to form a nucleic acid separating vessel assembly.
(3) A method for separating nucleic acid from a biological sample comprising the steps of: charging samples containing nucleic acid into the nucleic acid separating vessel described in the above (1) or the nucleic acid separating vessel assembly described in the above (2); combining the nucleic acid in the sample with the silicon oxide composite particle structural body of the nucleic acid separating vessel; washing the silicon oxide composite particle structural body, whereon the nucleic acid is combined; and releasing and recovering the nucleic acid from the silicon oxide composite particle structural body.
(4) Preferably, the particle which becomes a nucleus described in the above (1) is a resin particle.
(5) As described previously, the diameters of the silicon oxide particles and the resin particle can be set arbitrary. However, in view of forming the composite structure, the diameter of the silicon oxide group particle is preferably smaller than {fraction (1/10)} of the resin particle which becomes a nucleus described in the above (1). In accordance with the size of the micro plate, the maximum diameter of the resin particle is restricted to substantially 9 mm, and in view of problems in handling the particles such as scattering at mixing the particles, the minimum diameter of the silicon oxide particle is preferably at least 1 nm, that is, {fraction (1/9,000,000)} of the diameter of the resin particle.
(6) A method for manufacturing the nucleic acid separating vessel for separating nucleic acid from a biological sample comprising the steps of: forming a silicon oxide composite particle by making silicon oxide group particles form a composite with surface of a particle, which is larger than the silicon oxide group particle and becomes a nucleus; packing the plural silicon oxide composite particles into a die of a designated shape; forming the silicon oxide composite particle structural body having the designated shape by heating the die to melt and combine the plural silicon oxide composite particles each other three dimensionally; and assembling the silicon oxide composite particle structural body having the designated shape with a main body of the nucleic acid separating vessel as a nucleic acid separating portion.
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Proc. Natl. Acad. Sci. USA, vol. 76, No. 2, Feb. 1979, “Preparative and analytical purification of DNA from agarose”, B. Vogelstein et al, pp. 615-619.
Endo Yoshishige
Fukuzono Shinichi
Ikeda Yukiko
Yokoyama Tetsuo
Hitachi , Ltd.
Mattingly Stanger & Malur, P.C.
Redding David A.
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