Insect, insect hemolymph and assembly active substance thereof and application

A technology of hemolymph and active substances, applied in the field of collection and stable preservation of insect hemolymph

Inactive Publication Date: 2014-05-14
SHENYANG PHARMA UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0011] At present, there is no research on the components and biological activities of the hemolymph phenol oxidase proactivation system and Toll signal transmission system in Lepidoptera and Cephalidae insects at home and abroad

Method used

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  • Insect, insect hemolymph and assembly active substance thereof and application
  • Insect, insect hemolymph and assembly active substance thereof and application
  • Insect, insect hemolymph and assembly active substance thereof and application

Examples

Experimental program
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Effect test

Embodiment 1

[0101] Collection and storage of hemolymph

[0102] This embodiment enables those skilled in the art to understand the present invention more comprehensively, but does not limit the scope of the appended claims of the present invention in any way.

[0103] Use sterile water (distilled water or deionized water) to repeatedly wash the insects 3-4 times by means of rinsing; after cleaning, dry the body surface liquid and then use 75% alcohol to carry out body surface disinfection. Anesthetize the insects by placing them in an ice bath until unresponsive. Collect and store hemolymph separately according to the following conditions:

[0104] (1) Using the perfusion method, the anticoagulant buffer solution (15mmol.L -1 NaCl; 136mmol.L -1 Sodium citrate; 26mmol.L -1 Citric acid; 20mmol.L -1 EDTA, pH 4.0) was pre-injected into the body cavity of insect larvae, and the hind legs of the insect were removed or the abdomen of the insect was torn open to collect the hemolymph. After...

Embodiment 2

[0112] Response of pro-hemolymph phenoloxidase activation system to microbial infection in insects

[0113] Utilizing the basic procedures, principles, conditions, requirements, methods, etc. and their characteristics of insect detection of microorganisms, the experiment was carried out with tussah silkworm, the representative of Lepidoptera insects.

[0114] Take eight groups of tussah larvae, 10 in each group. Escherichia coli (Gram-negative bacteria) suspension, Staphylococcus aureus (Gram-positive bacteria) suspension, Candida albicans (fungus) suspension, peptidoglycan solution, β-1,3-glucan Sugar solution, lipopolysaccharide solution and lipoteichoic acid solution were injected into tussah silkworm larvae respectively, and physiological saline was used as blank control to observe the color change of tussah silkworm larvae's body surface.

[0115] The result is as image 3 As shown, the sample groups all caused the melanization and death of the tussah larvae body surface;...

Embodiment 3

[0118] Activation of prophenoloxidase in hemolymph by microbe-associated molecular patterns in vitro

[0119] The basic procedures, principles, conditions, requirements, methods, etc. and their characteristics of detecting microbial-related molecular patterns in insect lymph collection fluid were tested with tussah silkworm, a representative of Lepidoptera insects.

[0120] Microbe-related molecular patterns—lipopolysaccharide, peptidoglycan, mannan, lipoteichoic acid and β-1,3-glucan—were mixed with insect hemolymph collection fluid separately or simultaneously, and normal saline was used as a blank control. After incubation at 25°C and pH 5, the absorbance at 490nm was measured with dopa as the detection substrate to investigate the effect of microbial-related molecular patterns on the activity of phenoloxidase in the hemolymph collection fluid.

[0121] Experimental results such as Figure 4 As shown, the experimental group (column Figure 2-9 ) in the insect hemolymph co...

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Abstract

The invention relates to the field of biotechnology, and particularly relates to the bioactivity of the insect, insect hemolymph and assembly active substance thereof in activating a prophenoloxidase activation system or/and Toll signal transmission path system and an application of the insect, insect hemolymph and assembly active substance thereof in microorganism and related molecular mode detection thereof, preparation of the assembly active substance of prophenoloxidase activation system and separation and purification of antibacterial peptide. The invention also provides an insect hemolymph collection method and the storage method and conditions of the hemolymph; under the conditions, the stability of the insect hemolymph is better, and the functions of the insect hemolymph can be better played.

Description

Technical field: [0001] The invention relates to the field of biotechnology, and relates to a method for collecting and stably preserving insect hemolymph and its application. Specifically, the present invention relates to a method for collecting insect hemolymph containing the prophenoloxidase activation system or / and Toll signal transmission pathway (system) components, aiming at the prophenoloxidase activation system or / and Toll signal transmission pathway ( System) bioactive and stable insect hemolymph collection method and its hemolymph storage method and conditions, the biological activity of the prophenoloxidase activation system using insects and their hemolymph and the biological activity of the Toll signal transmission system, insect hemolymph phenol oxidation Active components of zymogen activation system and Toll signal transmission system and their reassembled biological activities, pattern recognition receptors obtained from insect hemolymph, cascade system serin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/07C12Q1/04C12Q1/26C07K14/435
Inventor 张嵘赵明沂张景海吴春福
Owner SHENYANG PHARMA UNIVERSITY
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