Preparation method of flow cytometry sample for chrysanthemum

A flow cytometry and sample preparation technology, applied in the preparation of test samples, etc., can solve the problems of complex origin and genetic background, and the inability to obtain ideal results, so as to improve accuracy, protect DNA from degradation, and prepare The effect of process reduction

Inactive Publication Date: 2015-02-25
BEIJING AGRO BIOTECH RES CENT
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Problems solved by technology

[0005] Chrysanthemum is rich in varieties, widely distributed, and has various shapes. Compared with many other plants, its origin and genetic background are much more complicated. As a result, some varieties of chrysanthemums cannot get ideal results when they are treated with conventional cell lysis buffer. Therefore, it is necessary to improve the cell lysis buffer and staining solution according to the special morphological structure and physiological characteristics of chrysanthemum, and the finally prepared samples are suitable for flow cytometry analysis

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  • Preparation method of flow cytometry sample for chrysanthemum
  • Preparation method of flow cytometry sample for chrysanthemum
  • Preparation method of flow cytometry sample for chrysanthemum

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preparation example Construction

[0032] A flow cytometry sample preparation method applicable to chrysanthemum, comprising the following steps:

[0033] The first step, lysing step: lysing, filtering, and centrifuging the chrysanthemum tissue sample to obtain chrysanthemum cell nuclei to be stained.

[0034] Specifically, this step specifically includes the following operations:

[0035] A. Get the chrysanthemum leaves, put them into a petri dish (preferably 6 cm in diameter) filled with pre-cooled cell lysis buffer, and cut the chrysanthemum leaves into pieces within 1 min with a sharp blade;

[0036] The types of chrysanthemums are: chamomile, heterochromia chrysanthemum, Asian chrysanthemum, Maohua chrysanthemum, Korean chrysanthemum, Japanese daisy, artemisia japonica, and purple wild chrysanthemum; the chrysanthemum leaves are selected from the new young leaves of the upper part of the unflowered chrysanthemum;

[0037] The mass volume (w / v) ratio of chrysanthemum leaves and cell lysis buffer is 1-10:10...

Embodiment 1

[0071] A flow cytometry sample preparation method applicable to chrysanthemum, comprising the following steps:

[0072] (1), take 50 mg of chrysanthemum leaves, put them into a petri dish dripped with 1 ml of pre-cooled cell lysis buffer, and cut the chrysanthemum leaves into pieces within 1 min with a Gillette double-sided blade;

[0073] The types of chrysanthemums are: chamomile, heterochromia chrysanthemum, Asian chrysanthemum, Maohua chrysanthemum, Korean chrysanthemum, Japanese daisy, artemisia japonica, and purple wild chrysanthemum; the chrysanthemum leaves are selected from the new young leaves of the upper part of the unflowered chrysanthemum;

[0074] The components of the cell lysis buffer are:

[0075] 15mM MOPS, 2mM Na 2 EDTA, 0.5mM spermine.4HCL, 80mM KCl, 20mM NaCl, 0.1% (v / v) TritonX-100, 1% (w / v) pvp-10, 0.1% (v / v) β - mercaptoethanol, pH 7.5;

[0076] (2), then add 1 ml of pre-cooled cell lysis buffer to the fragments to release the nuclei to obtain a nuc...

Embodiment 2

[0106] A flow cytometry sample preparation method applicable to chrysanthemum, comprising the following steps:

[0107] (1), take 50 mg of chrysanthemum leaves, put them into a petri dish dripped with 1 ml of pre-cooled cell lysis buffer, and cut the chrysanthemum leaves into pieces within 1 min with a Gillette double-sided blade;

[0108] The types of chrysanthemums are: chamomile, heterochromia chrysanthemum, Asian chrysanthemum, Maohua chrysanthemum, Korean chrysanthemum, Japanese daisy, artemisia japonica, and purple wild chrysanthemum; the chrysanthemum leaves are selected from the new young leaves of the upper part of the unflowered chrysanthemum;

[0109] The components of the cell lysis buffer are:

[0110] 15mM MOPS, 2mM Na 2 EDTA, 0.5mM spermine.4HCL, 80mM KCl, 20mM NaCl, 1% (v / v) TritonX-100, 0% (w / v) pvp-10, 0.2% (v / v) β - mercaptoethanol, pH 7.5;

[0111] (2), then add 1 ml of pre-cooled cell lysis buffer to the fragments to release the nuclei to obtain a nucle...

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Abstract

The invention belongs to the field of the detection on flow cytometry, and provides a preparation method of a flow cytometry sample for chrysanthemum. The method comprises the following steps: carrying out a splitting decomposition step, namely, carrying out the splitting decomposition on a chrysanthemum sample, filtering and centrifuging to obtain a cell nucleus to be dyed; and carrying out a dyeing step, namely, dyeing the cell nucleus to be to be dyed to obtain the dyed cell nucleus. According to the flow cytometry analysis, an appropriate cell lysis buffer solution is adopted, so that cytoplasmic fragments remained in the complete cell nucleus of the chrysanthemum can be effectively eliminated, the stability of the cell nucleus in suspension liquid can be maintained, and the agglutination of the cell nucleus in the suspension liquid can be prevented; DNA can be protected from degradation; an appropriate environment is provided for the exclusive nucleus DNA chemical staining, and the adverse impact of the cytoplasm component on the dyeing can be effectively reduced; by optimizing the sample preparation flow, the disturbance peak in the computer testing can be effectively reduced, and the precision, accuracy and stability of a detection result can be improved.

Description

technical field [0001] The invention belongs to the field of flow cytometry detection, and particularly relates to a method for preparing a flow cytometry sample suitable for chrysanthemums. The method prepares a suitable reaction solution such as a cell lysis buffer for chrysanthemums to obtain a suitable sample for flow cytometry. Chrysanthemum samples analyzed by cytometry. Background technique [0002] Chrysanthemum (chrysanthemum morifolium) is Compositae (compositae) Chrysanthemum (chrysanthemum) plant, originates in my country, is one of Chinese tradition ten famous flowers and world four big cut flowers, is also the city flower of Beijing. my country is the origin center of cultivated chrysanthemum and the distribution center of Chrysanthemum germplasm resources (Li Xinlei, 2004). Wild resources and cultivated varieties are very rich, and its petal and flower patterns vary greatly. During the cultivation and application history of more than 1,600 years, a large numb...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/28
Inventor 罗昌陈东亮黄丛林程曦梁宏霞苏国辉黄敦辉
Owner BEIJING AGRO BIOTECH RES CENT
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