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Fluorescence in-situ hybridization method for endosymbiotic bacteria in insect ovary

A technique of fluorescence in situ hybridization and symbiotic bacteria, which is applied to the field of in situ hybridization of symbiotic bacteria in insect ovaries, can solve the problems of time-consuming, labor-intensive, complicated operation, etc., and achieve the effects of simple operation, less chemical reagents, and clear staining results.

Inactive Publication Date: 2018-06-19
INST OF COTTON RES CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The traditional fluorescent in situ hybridization staining method for insect tissue generally requires freezing and sectioning insect tissue, which is cumbersome, time-consuming and labor-intensive in operation, and it takes 2-3 days to complete an experiment

Method used

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  • Fluorescence in-situ hybridization method for endosymbiotic bacteria in insect ovary
  • Fluorescence in-situ hybridization method for endosymbiotic bacteria in insect ovary
  • Fluorescence in-situ hybridization method for endosymbiotic bacteria in insect ovary

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Fluorescence in situ hybridization of symbiotic bacteria Wolbachia in the ovary of Bemisia tabaci

[0046] Fluorescent in situ hybridization for endosymbiont Wolbachia in whitefly ovary with fluorescent probe

[0047] W2: Cy5-CTTCTGTGAGTACCGTCATTATC.

[0048] Including the following steps:

[0049] (1) Add PBS buffer solution to the petri dish, and put the adult female insects to be dissected into the PBS buffer solution;

[0050] (2) Under the stereoscopic anatomical microscope, fix the insect chest web with an insect needle, cut open the abdomen of the insect with a dissecting needle, pull out all the ovaries, and remove debris such as residual shells of the insect body;

[0051] (3) Add 3-5 drops of PBS buffer in the center of the slide, and pick the ovary in the petri dish into the PBS buffer on the slide with a fine brush;

[0052] (4) Use filter paper to suck off the PBS buffer around the ovary on the glass slide, and immediately add 200ul of insect tissue fixati...

Embodiment 2

[0062] Fluorescence in situ hybridization of symbiotic bacteria Portiera in whitefly whitefly ovary was carried out with the fluorescent probe BTP1:Cy3-TGTCAGTGTCAGCCCAGAAG.

[0063] The hybridization method was the same as in Example 1 except that the fluorescent probes used were different. see results image 3 .

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Abstract

The invention relates to a fluorescence in-situ hybridization method for endosymbiotic bacteria in an insect ovary. The method comprises following steps: the insect ovary is dissected under a microscope, the dissected insect ovary is transferred to a glass slide, an insect tissue stationary liquid is added for fixation, and a destaining solution is added for destaining; pre-hybridization and hybridization staining are performed on a specimen, and an eluant is added for elution after hybridization is completed; finally, a sample is sealed with glycerin, and observation and photographing are performed under a confocal microscope. By means of the method, a good cell staining effect can be kept, the operation can be simplified, time can be shortened, reagents can be saved and the cost can be reduced. Several endosymbiotic bacteria in the ovary can be subjected to multiple staining simultaneously, a good staining effect is obtained, and the method is simple, convenient, rapid and economical.

Description

technical field [0001] The invention belongs to the field of biological technology, and particularly relates to an in-situ hybridization method of symbiotic bacteria in insect ovaries. Background technique [0002] Insect endosymbionts are a type of symbiotic microorganisms that live in insects and co-evolve with their hosts. They participate in the nutritional metabolism of the host, provide nutrients for the host, enhance the host's defense against natural enemies, regulate the host's heat resistance, and enhance the host's resistance. Medicinal properties and other functions. The infection of endosymbionts has an important impact on the reproduction, survival and adaptability of the host to the environment, and is considered to be an important regulatory factor in the growth and development of insects. The role of endosymbionts is closely related to their distribution in insect tissues. Some symbionts are mainly distributed in ovaries and testes, and participate in the r...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/689C12Q1/6841
CPCC12Q1/6841C12Q1/689C12Q2563/107
Inventor 胡红岩马艳任相亮姜伟丽马小艳马亚杰王丹
Owner INST OF COTTON RES CHINESE ACAD OF AGRI SCI
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