A kind of in situ hybridization method of Lagerstroemia genus chromosome

An in situ hybridization and chromosome technology, applied in biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of difficult to identify chromosomes, blank, difficult to dye, etc., and achieve high hybridization specificity

Inactive Publication Date: 2016-01-20
BEIJING FORESTRY UNIVERSITY
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

Most of the plants in this genus have small and medium-sized chromosomes, and they are not easy to stain. It is difficult to identify their chromosomes by classical cytological methods, which limits the in-depth understanding of the molecular cytogenetic mechanism of Lagerstroemia indica
So far, there is no report on the application of in situ hybridization technology in Lagerstroemia genus, and the research on its genome structure and molecular cytogenetics is still blank.

Method used

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  • A kind of in situ hybridization method of Lagerstroemia genus chromosome
  • A kind of in situ hybridization method of Lagerstroemia genus chromosome

Examples

Experimental program
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Effect test

Embodiment 1

[0049] 1. Preparation of chromosome slide specimens:

[0050]At 9:00 in the morning, take the stem tip part of the innermost length of the shoot tip growth point of the annual branch of Lagerstroemia urophyllum with a length of 0.5 cm. Put them directly into the fixative solution (by volume, absolute ethanol: glacial acetic acid (analytical grade): chloroform (analytical grade) = 5:3:2) and fix in a -20°C refrigerator for 18 hours. Rinse and wash the fixative in distilled water, transfer to 0.075mol / L KCl solution and hypotonic for 30 minutes; rinse the growth point of the shoot tip after the pre-hypotonic treatment with distilled water, dry it with absorbent paper, and put it into the mixed enzyme solution Enzymolysis was performed in a water bath at 37°C for 4 hours, and the mixed enzyme solution was prepared from 2.5% cellulase: 2.5% pectinase: 0.01% proteinase K at a volume ratio of 2:1:3. The slides were prepared by the flame drying method, and the microscopic examinatio...

Embodiment 2

[0066] The difference from Example 1 is that the material is Yakushima crape myrtle. The steps and signal observation are the same as in Example 1.

[0067] The results show that from figure 1 and figure 2 As can be seen in , there are 2 signal points (marked by arrows) when the Dig-labeled maize 45S rDNA probe hybridizes to the Lagerstroemia genus chromosome preparation. The background interference is weak, and the sensitivity and accuracy are high, so that the in situ hybridization technique can be better applied to the chromosome detection of small chromosome plants such as Lagerstroemia indica, and provides a new way and method for the in-depth study of the chromosomes of Lagerstroemia genus plants.

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Abstract

The invention relates to a lagerstroemia plant chromosomal in-situ hybridization method. The method comprises seven steps such as sheet making, probe preparing, pretreating before hybridization, hybridization solution preparing, probe denaturating, chromosome denaturating, hybridizing and detecting. A lagerstroemia plant metaphase chromosome serves as a target deoxyribose nucleic acid (DNA) and maize 45SrDNA as a probe, and the maize 45SrDNA probe is clearly located on the lagerstroemia plant metaphase chromosome. By utilizing the hybridization method, background interferences can be effectively eliminated, the detection sensitivity and accuracy are improved, the hybridization method can be better applied to the chromosome detection of small chromosome plants such as lagerstroemia and the like, and a new way and a method are provided for the further research of lagerstroemia plant chromosomes.

Description

technical field [0001] The invention relates to the field of biological detection, in particular to an in situ hybridization method for chromosomes of Lagerstroemia genus plants. Background technique [0002] Molecular in situ hybridization (Insituhybridization, referred to as ISH) is to use DNA fragments labeled with biotin and other markers as probes to carry out molecular hybridization with chromosomal DNA, and directly detect the existence of DNA fragments complementary to the probe under a microscope parts. The physical positioning of rDNA on chromosomes by fluorescence in situ hybridization technology can not only provide a stable and effective cytologically identifiable chromosome marker for karyotype analysis, but also study the evolutionary relationship between plant species and clarify the structural variation of chromosomes and other important genetic issues. At present, rDNA has been extensively physically located in the genomes of important model crops and com...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 潘会堂杨冰洁张启翔蔡明程堂仁王佳孙明陈晶鑫
Owner BEIJING FORESTRY UNIVERSITY
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