Method for separating and purifying insect pathogenic fungoid thallus from infected insect haemolymph

A technology for the separation and purification of pathogenic fungi, applied in biochemical equipment and methods, fungi, microorganism measurement/inspection, etc., can solve the problems of time-consuming, difficult detection of pathogenic fungal genes, laborious and other problems

Inactive Publication Date: 2006-07-26
CHONGQING UNIV +1
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Problems solved by technology

[0002] At present, the research on the pathogenic gene of entomopathogenic fungi is mainly to establish the induced expression sequence tag library (hereinafter referred to as the EST library) under different in vitro conditions, such as by establishing Metarhizium anisopliae (broad-spectrum bacterial strain) in different in vitro conditions. Under the induced EST library, analyze a large number of EST sequences, in order to use a large number of induced EST sequences under different in vitro conditions to analyze the gene expression of pathogenic fungi after invading host insects, etc., but this method is time-consuming, laborious, and difficult. Detection of pathogenic fungal genes specifically expressed under in vivo conditions

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  • Method for separating and purifying insect pathogenic fungoid thallus from infected insect haemolymph
  • Method for separating and purifying insect pathogenic fungoid thallus from infected insect haemolymph
  • Method for separating and purifying insect pathogenic fungoid thallus from infected insect haemolymph

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Embodiment Construction

[0028] A method for separating and purifying entomopathogenic fungal cells from hemolymph of susceptible insects, characterized in that: hemocytes of susceptible insects are lysed by enzyme treatment and their DNA and RNA are degraded by enzyme treatment, followed by separation and precipitation. The pathogenic fungal cells were purified and obtained; during the whole enzymatic hydrolysis process, PCR was used to detect the conserved sequence monitoring, and finally PCR and RT-PCR were used to detect the conserved sequences to identify the entomopathogenic fungal cells.

[0029] In a word, the theoretical basis of the present invention is designed by utilizing the structural differences between animal cells and fungal cells, because the outermost layer of animal cells is wrapped by cell membranes, and the cell membranes are composed of double-layer phospholipids and membrane proteins. Therefore, proteinase K is used to specifically degrade proteins to degrade membrane proteins ...

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Abstract

This invention discloses a method for segregation purifying insect pathogenic fungi thalline from suscepted insect bloodlympha. It is prepared through steps as follows: deal with blood cell of suscepted insects with a series of enzymes to destroys and removes their germ plasms; finally seperate and purify to get the products. Adopt the achieved pathogenic fungi thalline to extract DNA and mRNA, which has no insect germ plasm pollution according to PCR and RT-PCR verification. The DNA and mRNA extracted can be used to construct gene library, it also lays solid foundation for further uncovering entomogenous fungi invading mechanism.

Description

technical field [0001] The invention relates to a method for separating and purifying entomopathogenic fungal cells from hemolymph of susceptible insects. Background technique [0002] At present, the research on the pathogenic gene of entomopathogenic fungi is mainly to establish the induced expression sequence tag library (hereinafter referred to as the EST library) under different in vitro conditions, such as by establishing Metarhizium anisopliae (broad-spectrum bacterial strain) in different in vitro conditions. Under the induced EST library, analyze a large number of EST sequences, in order to use a large number of induced EST sequences under different in vitro conditions to analyze the gene expression of pathogenic fungi after invading host insects, etc., but this method is time-consuming, laborious, and difficult. Detection of pathogenic fungal genes specifically expressed under in vivo conditions. [0003] The separation and purification of pathogenic fungal cells ...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12S3/22C12Q1/68
Inventor 夏玉先张沧桑王中康殷幼平彭国雄
Owner CHONGQING UNIV
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