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Method for separating chromosomes in plant cells by using fluorescence in-situ hybridization

A fluorescence in situ hybridization and plant cell technology, applied in DNA preparation, chemical library, combinatorial chemistry, etc., can solve the problems of immature plant chromosome banding technology and application limitations, avoid chemical treatment, broaden the scope, and prevent damage effect

Inactive Publication Date: 2009-11-18
INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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Problems solved by technology

However, it is still difficult to produce chromosome specimens with clear banding and suitable for micro-separation, and many plant chromosome banding techniques are immature, so its application is limited

Method used

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  • Method for separating chromosomes in plant cells by using fluorescence in-situ hybridization
  • Method for separating chromosomes in plant cells by using fluorescence in-situ hybridization
  • Method for separating chromosomes in plant cells by using fluorescence in-situ hybridization

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Embodiment 1

[0019] Embodiment 1, the microseparation technique of wheat chromosome

[0020] 1. Chromosome production

[0021] Wheat seeds germinated on a petri dish with double-layer filter paper at 23°C, and treated at 4°C for 24 hours after dew blanching, then transferred to 23°C for constant temperature cultivation for 24 hours, cut the roots of germinated seeds, and treated them under ice water at 0°C for 36 hours. Hours, after absorbing the water on the surface of the root tip, place it in fresh Carnoy's fixative solution (absolute ethanol: glacial acetic acid = 3: 1) and fix it for 10 minutes, then transfer it to 70% ethanol (volume percentage), Store at 4°C until use. Get the fixed root tip, rinse it with sterile water, cut the root tip under aseptic conditions and place it in 2% cellulase (R-10 Yakult.) and 2% pectinase (Y-23 Yakult.) 1: In the mixed solution prepared in 1, enzymatically hydrolyze at 37°C for 3 hours, gently rinse twice with sterile water, stain with 1% acetic a...

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Abstract

The invention discloses a method for separating chromosomes in plant cells by using fluorescence in-situ hybridization. The method for separating the chromosomes in the plant cells comprises the following steps of: the fluorescence in-situ hybridization is carried out on the plant cells, the single chromosome of which is picked; by comparing with the corresponding fluorescence in-situ hybridization map of the normal plant cells, the picked single chromosome is defined to be which chromosome, thus separating a purpose chromosome. The method has the advantages of being capable of separating the chromosomes with similar types and specific bands, widening the separating range of the chromosomes, avoiding the unnecessary chemical processing in the manufacture process of the chromosome specimens, preventing the DNA of the chromosomes from being damaged, guaranteeing the completeness of the separated DNA. A microclone library of the whole chromosomes of the wheat can be obtained by applying the invention.

Description

technical field [0001] The invention relates to a method for separating chromosomes in plant cells by using fluorescence in situ hybridization. Background technique [0002] Chromosome microsegregation refers to a technique in which chromosomes are separated or cut under a microscope to obtain specific chromosomes or chromosome fragments. It is the basis of chromosome microcloning and a powerful tool for genetics research. At present, there are three main methods for chromosome microseparation and cutting, namely, glass needle method, laser cutting method, and flow cytometry method. Among them, the glass needle method uses an inverted microscope to directly cut the target chromosome with a fine glass with a tip diameter of no more than 0.5 μm. This method is favored by most researchers because of its simplicity, convenience and low cost. [0003] Due to the special structure of plant cells and chromosomes, the research on micro-separation technology of plant chromosomes st...

Claims

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Application Information

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IPC IPC(8): C12N15/10C40B50/06
Inventor 郑琪李振声胡赞民李滨李宏伟
Owner INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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