Method for preparing conventional slide of chromosomes from grapevine root tips

A chromosome and grape root technology, applied in the field of cytogenetics, can solve the problems of the small number of grape chromosomes and the limitations of chromosome research, and achieve the effects of reducing cytoplasmic viscosity, large dispersion space, and easy counting

Inactive Publication Date: 2017-11-24
SHANXI AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years, the research on grape genetics and breeding has developed rapidly. However, due to the characteristics of small and large number of grape chromosomes

Method used

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  • Method for preparing conventional slide of chromosomes from grapevine root tips
  • Method for preparing conventional slide of chromosomes from grapevine root tips
  • Method for preparing conventional slide of chromosomes from grapevine root tips

Examples

Experimental program
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Effect test

example 1

[0032] The diploid Lihongbao grape root tip dyeing chromosome compression method comprises the steps in the following order:

[0033] (1) When cultivating materials, choose robust annual Lihongbao branches during winter shearing. After three months of winter sand storage, take out the branches and cut the branches into cuttings with a length of 10-12cm, leaving 2-3 buds for each cutting. , when cutting the cuttings, the distance between the upper notch and the upper bud is 1.5~2 cm, cut horizontally, the lower notch is close to the cut part, cut obliquely, completely immerse the cuttings in clean water for 24 hours, until the cut section is bright green, Take out and drain, and immerse the base downward in 100 mg / L naphthalene acetic acid solution for 24 hours to promote rooting. Put the cuttings into conical flasks for cultivation at a water depth of 2-4 cm, and place them in a light incubator with a light intensity of 2000 Lx, light for 12 h, culture at a constant temperatur...

example 2

[0042] The triploid Nyahei root tip chromosome compression method comprises the steps in the following order:

[0043] Steps 1 and 2 are the same as in Example 1.

[0044] Step 3: Fix the pretreated Xiahei root tip material with distilled water for three times, and then place it in Carnot’s fixative (V absolute ethanol: V glacial acetic acid=3:1, Carnot’s fixative is prepared and used now), Fix at 4°C for 36 h.

[0045] Step 4 Dissociation Take out the fixed Xiahei root tip, wash it three times with distilled water, put it in a 2 mL centrifuge tube, then add 3.5% mixed enzyme solution (same as in Example 1) to the centrifuge tube, mix the enzyme solution and the root tip The volume ratio of 30:1, 37 ℃ constant temperature enzymatic hydrolysis for 25 minutes, then rinsed with distilled water 2~3 times, placed in distilled water for later use.

[0046] Steps 5, 6 and 7 are the same as in Example 1. Microscopic examination results and photomicrographs see figure 2 .

example 3

[0048] The tetraploid Jingya root tip chromosome compression method includes the steps in the following order:

[0049] Except that the fixed time in step 3 was 48 h, and the dissociation and enzymatic hydrolysis in step 4 were 30 min, the rest of the steps were the same as in Example 2. Microscopic examination results and photomicrographs see image 3 .

[0050] The results of the above implementation and application on three representative grape varieties show that this method is an effective method for conventional grape chromosome compression, and the compression method is simple and convenient to operate, the chromosomes are evenly dispersed, the background is light, and the number of chromosomes is easy to obtain. Counting, the number of chromosomes obtained by this method has high accuracy.

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Abstract

The invention discloses a method for preparing a conventional slide of chromosomes from grapevine root tips. The method comprises the following steps: material culture, pretreatment, immobilization, preservation, dissociation, dyeing, slide preparation and microscopic examination. According to the invention, rooting of grapevine twigs is easy, and root tips are moderately thick and facilitate preparation of a chromosome slide; the prepared slide is flat in specimen cells and dispersed in chromosomes; pretreatment with a saturated p-dichlorobenzene solution can improve the success rate of slide preparation, enables chromosomes to be shortened and thickened, reduces the viscosity of cytoplasm and allows more metaphase chromosomes to be obtained; and a mixed enzyme solution with a concentration of 3.5% is used for enzymatic hydrolysis, so cell walls are decomposition, pectic substances among meristematic cells are decomposed, cells are well dispersed, and chromosomes are large in dispersion space and uniform in dispersion and are thus easy to count. The preparation method of the invention has the advantages of easiness, quick preparation, low preparation cost and easy operation, and obtained cells are good in division phase and high in accuracy, which are of great significance to understanding of the genetic background of grapes, the selection of parents in breeding and post-identification.

Description

technical field [0001] The invention belongs to the field of cytogenetics, and is one of the ploidy identification methods of grape plants, in particular to a method for routine pressing of grape root tip chromosomes. Background technique [0002] Grape( Vitis vinifera L.) is a woody vine of the genus Vitis vinifera, and is one of the oldest fruit tree species in China. Whether in fresh food or brewing and processing, it is deeply loved by people, and it is the unremitting pursuit of grape breeding scientists to continuously cultivate new and excellent grape varieties. [0003] Chromosomal ploidy identification has important guiding significance for improving grape traits, improving breeding efficiency, and cultivating fine varieties. The observation of chromosome number, shape and structure is of great significance to the research of genetic breeding. Changes in the structure and number of chromosomes will lead to the variation of organisms. Therefore, chromosome countin...

Claims

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Application Information

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IPC IPC(8): G01N1/28G01N1/30
CPCG01N1/2813G01N1/30G01N2001/302
Inventor 纪薇王静波赵旗峰董志刚王敏
Owner SHANXI AGRI UNIV
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