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Method for quickly and effectively transfecting mosquito by using wolbachia

An effective and fast technology, applied in animal husbandry and other fields, can solve the problem of losing precious primary generations, and achieve the effect of saving time and energy, simple operation and easy access

Active Publication Date: 2012-02-15
GUANGZHOU WOLBAKI BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But this method is inappropriate, because in theory, the positive in the primary (G0) does not represent the success of the transfection, and at the same time, this method discards the negative strains in the primary (G0), which will lose a lot The precious primary generation (G0), because some positive strains exist in the offspring (G1) of these discarded negative primary generations (G0)

Method used

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  • Method for quickly and effectively transfecting mosquito by using wolbachia
  • Method for quickly and effectively transfecting mosquito by using wolbachia
  • Method for quickly and effectively transfecting mosquito by using wolbachia

Examples

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Effect test

Embodiment 1

[0026]1. Acquisition of Wolbachia: Collect the ovaries of Aedes albopictus containing Wolbachia by dissecting them, and dissect multiple mosquitoes at one time to collect their ovaries. The ovaries collected at one time are very important. About 50 mg was collected and placed in 1 ml of SPG buffer. The ovarian tissue was crushed with a Dounce Tissue Grinder, and the ovarian tissue was ground 10 times by the Dounce Tissue Grinder, 0.5 min each time, and the ground tissue was washed with 0.5 ml of SPG buffer collect. The collected liquid was then centrifuged at 300 g for 5 minutes in a centrifuge to remove some tissue fragments. Transfer the supernatant to a new centrifuge tube and centrifuge again at 12000g for 10 minutes. The supernatant was removed, and the sediment in the centrifuge tube contained purified Wolbachia. After dissolving the sediment in the centrifuge tube with 50 microliters of SPG buffer solution, centrifuge again at 300g for 5 minutes to further remove the...

Embodiment 2

[0030] 1. Acquisition of Wolbachia: Collect the ovaries of Aedes albopictus containing Wolbachia by dissecting them, and dissect multiple Aedes albopictus at one time to collect their ovaries. The ovary of Aedes albopictus weighs about 50 mg and is collected in 1 ml of SPG buffer. The ovarian tissue was crushed with a Dounce Tissue Grinder, and the ovarian tissue was ground 10 times by the Dounce Tissue Grinder, 0.5 min each time, and the ground tissue was washed with 0.5 ml of SPG buffer collect. The collected liquid was then centrifuged at 100g in a centrifuge for 10 minutes to remove some tissue fragments. Transfer the supernatant to a new centrifuge tube and centrifuge again at 15000g for 5 minutes. The supernatant was removed, and the sediment in the centrifuge tube contained purified Wolbachia. After dissolving the sediment in the centrifuge tube with 50 microliters of SPG buffer solution, centrifuge again at 100g for 10 minutes to further remove the remaining tissue ...

Embodiment 3

[0034] 1. The acquisition of Wolbachia (Wolbachia) is as follows: the Culex mosquito (Cx.pipiens molestus) containing Wolbachia (Wolbachia) is collected by dissecting its ovaries, and a plurality of Culex mosquitoes are dissected at one time to collect its ovaries. Culex ovaries collected at one time weigh about 50 mg, and placed in 1 ml of SPG buffer after collection. The ovarian tissue was crushed with a Dounce Tissue Grinder, and the ovarian tissue was ground 10 times by the Dounce Tissue Grinder, 0.5 min each time, and the ground tissue was washed with 0.5 ml of SPG buffer collect. The collected liquid was then centrifuged at 1000g in a centrifuge for 1 minute to remove some tissue fragments. Transfer the supernatant to a new centrifuge tube and centrifuge again at 5000g for 20 minutes. The supernatant was removed, and the sediment in the centrifuge tube contained purified Wolbachia. After dissolving the sediment in the centrifuge tube with 50 microliters of SPG buffer ...

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Abstract

The invention discloses a method for quickly and effectively transfecting mosquito by using wolbachia. According to the method, an insects ovarian tissue (abundant sources and easiness for obtaining) containing the wolbachia is directly acquired by adopting a vivisection method and is homogenated; and the ovarian tissue is separated from the wolbachia by using a centrifugation method so as to obtain a purified usable wolbachia transfected by embryonic injection. The embryo of the mosquito is injected and transfected by the purified wolbachia through a conventional method; a primary female adult insect (G0) is obtained by egg culture; the obtained primary female adult insect is mated with a wild type male mosquito to generate first generation (G1) mosquito strains; and positive mosquito strains containing the wolbachia are screened from the first generation mosquito strains and become really and successfully-transfected mosquito strains. Therefore, the method for transfecting the mosquito by using the wolbachia is a quick and effective method. The realization of the invention is beneficial to quickly establishing the mosquito strains transfected with the wolbachia and further beneficial to large-scale application of the mosquito strains.

Description

Technical field: [0001] The invention belongs to the field of biotechnology, in particular to a method for quickly and effectively transfecting mosquitoes with Wolbachia. Background technique: [0002] Wolbachia is a symbiotic microorganism widely distributed in arthropods, and it may be the most abundant group of insect symbiotic microorganisms. It is distributed in Coleoptera, Diptera, Hemiptera, Homoptera, Hymenoptera, Lepidoptera and other insect species. It uses vertical transmission as its basic mode of transmission among host generations. It exists stably in the germ cells of the host, is transmitted to the offspring of the host through egg cells, and can regulate the reproductive activities of the host through various methods such as cytoplasmic incompatibility, feminization and andricide. These regulatory functions promote its widespread spread within the host population. [0003] Cytoplasmic incompatibility (CI) is the most common type of host reproductive behav...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01K67/033
Inventor 奚志勇朱俭
Owner GUANGZHOU WOLBAKI BIOTECH CO LTD
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