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Method and apparatus for sample preparation

A nucleic acid and individual technology, applied in biochemical equipment and methods, microbial measurement/inspection, fluorescence/phosphorescence, etc., can solve problems that are not suitable for digital counting, and achieve the prevention of sample loss and reduction of reaction efficiency, cost and Effects of labor saving, cost and labor reduction

Inactive Publication Date: 2008-10-01
HITACHI LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Recovery of generated DNA using solid beads is a good method, and it can be adequately used for purposes such as determining genome sequences using repeated DNA samples, but it is not suitable for digital counting

Method used

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  • Method and apparatus for sample preparation
  • Method and apparatus for sample preparation
  • Method and apparatus for sample preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] This example shows an example in which an emulsion containing agarose was prepared by stirring in oil.

[0062] exist figure 1 The basic concept of this method is shown in . By dividing the sample solution containing DNA molecules 1-3 to be analyzed into M micro-droplets 4-8 larger than the total number N of DNA molecules, micro-droplets 4, 5, 7, DNA into which DNA has entered are formed. Tiny droplets that did not enter6,8. The tiny droplets 4-8 are dispersed in the oil 10 in the reaction vessel 9 to form an emulsion 11 . After the emulsion containing the micro-droplets is subjected to an amplification reaction such as PCR, the presence or absence (amount) of the amplification product obtained in each micro-droplet is detected by fluorescence detection using an intercalator or the like, and is divided into 12 detections with fluorescence. The micro-droplets 13 of the amplified products from each DNA 1-3 and the micro-droplets 14 without the amplified products that c...

Embodiment 2

[0100] Embodiment 2: the shape of reaction vessel

[0101] In this embodiment, the shape of the reaction vessel is formed on the plate of the series of micro reaction wells separated from each other. use Figure 8-1 0 explains this embodiment. elephant Figure 8 Likewise, the plate 80 is provided with a plurality of wells 83 for collecting the respective minute droplets 81, 82. hole 83 as Figure 9 As shown, the 2-dimensional sequence constitutes the plate 80 . The tiny droplet directly enters the hole 83 and can be covered with a hydrophobic solvent 84 or the like, or can enter the hydrophobic solvent 84 in the hole 83 .

[0102] In this case, the hydrophobic solvent 84, in addition to the purpose of forming an emulsion, can also realize the function of preventing the evaporation of water in the reaction solution, keeping the shape of the tiny droplet as a spherical function, and preventing the gel and the surface of the container when the gel is taken out. The bonding ...

Embodiment 3

[0108] This example shows another example of the method for producing fine liquid droplets.

[0109] use Figure 11 This embodiment will be described. In this embodiment, the inkjet unit 100 is used to form minute droplets. The inkjet unit 100 is composed of a tank 101 for storing a solution for preparing fine liquid droplets 103 and a nozzle 102 for forming fine liquid droplets and ejecting them. By heating the reaction liquid in the nozzle instantaneously, a certain amount of the reaction liquid is ejected. The tiny liquid droplets 103 are arranged facing the container 105 so that they are sprayed or dropped directly into the hydrophobic solvent 104 . Micro-droplets 103 prepare emulsion 106 by spraying or falling into hydrophobic solvent 104 .

[0110] This embodiment is suitable for controlling the size and quantity of tiny liquid droplets, and is especially suitable for preparing liquid droplets from about 0.5 pl to about 10 pl (about 10 μm-30 μm in diameter). It is e...

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Abstract

A method of the present invention comprises fractionating a sample solution containing analyte DNA molecules into small droplets, wherein the number M of the droplets is greater than the total number N of the DNA molecules, subjecting an emulsion containing the droplets to, for example, PCR amplification, and detecting the presence or absence (amount) of an amplicon obtained in each droplet by fluorescent detection using an intercalator or the like.

Description

technical field [0001] The present invention relates to a sample preparation method used in gene analysis technology. More specifically, it relates to a sample preparation method for data analysis of mRNA contained in one cell or a method for simultaneously analyzing a plurality of target molecules one by one. Background technique [0002] With the completion of the decipherment of the human genome sequence, it is time to study various genome information in depth and make use of it. Genomic information is copied by mRNA and translated into protein. Gene expression profile analysis performed in this way is indispensable for detailed studies of life activities. Up to now, the mainstream analysis method is to extract mRNA from multiple cells, perform fluorescent labeling, let DNA probe array (DNA chip) function, and capture mRNA with a complementary sequence to the mRNA in the probe-labeled detection method . On the other hand, there is also a method of extracting mRNA from...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N21/64
CPCC12Q1/6806C12Q1/6851C12Q2527/143C12Q2563/173
Inventor 村川克二泷口纯代神原秀记
Owner HITACHI LTD
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