Pteromalus puparum venom protein PpVPG and application thereof

A technology of the venom glands of the golden bee and the golden bee, which is applied in the fields of genetic engineering, protein engineering, and molecular biology, and can solve problems such as ecological balance destruction, killing, and environmental pollution

Active Publication Date: 2019-12-20
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the long-term high-frequency use of chemical pesticides has caused problems such as pest resistance, beneficial biological killing, pesticide poisoning, excessive pesticide residues, r

Method used

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  • Pteromalus puparum venom protein PpVPG and application thereof
  • Pteromalus puparum venom protein PpVPG and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Embodiment 1, expression and purification of recombinant pET32a-PpVPG

[0044] 1.1. Extraction of RNA

[0045] Under a Leica MZ 16A microscope (Leica, Germany), the female bee was sterilized with 75% alcohol and wiped clean, its digestive tract was pulled out from the abdomen, and soaked in sterile 1 unit / μL RNase inhibitor ( TOYOBO, Osaka, Japan) in phosphate-buffered saline (PBS) droplets, collect the digestive tract in TRIzol reagent (Invitrogen, California, USA), and the RNA extraction method is as follows:

[0046] 1) Prepare to take multiple females of the chrysalis chrysalis on the third day of eclosion, collect them into a finger tube, freeze at -80°C for 10 minutes, take them out and insert them into an ice box. Take the parasitoid wasps and place them on the glass slide with PBS buffer, use dissecting forceps to remove the venom glands from the abdomen of the female bees, collect the dissected venom glands, add 1ml Trizol, shake vigorously, and let stand at r...

Embodiment 2

[0114] Example 2, Determination of Recombinant PpVPG Protein's Inhibitory Effect on PPO Activation in Pieris rapae

[0115] Put the hemolymph from the chrysalis chrysalis on ice into a 1.5ml pre-cooled centrifuge tube, and immediately put it on ice, centrifuge at 3300×g for 5 minutes at 4°C, transfer the supernatant to a new pre-cooled 1.5 ml centrifuge tube, remove the blood cells for the determination of the background of phenol oxidase.

[0116] Take 2 μl of hemolymph from each sample and add to 96-well plate wells containing 10 μl TBS (pH7.4) and 10 μl TBS containing 0.5 μg Micrococcus luteus (Micrococcus luteus), let stand at room temperature for 20 minutes, add 200 μl L dopa ( 2mM / L) was measured at a wavelength of 470nm for 30 minutes, and the unit U of phenol oxidase activity refers to the amount of 0.001OD that changes per minute. If the OD value of the sample wells with TBS and hemolymph is low, and the OD value of the sample wells of hemolymph with TBS and Micrococ...

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Abstract

The invention relates to the fields of molecular biology, gene engineering and protein engineering in particular to a protein PpVPG expressed in pteromalus puparum venom, and a coded nucleic acid sequence and application thereof. The invention discloses the pteromalus puparum venom protein PpVPG and a gene encoding the pteromalus puparum venom protein PpVPG. The pteromalus puparum venom protein PpVPG can be used for preparing venom protein PpVPG secreted by pteromalus puparum venom glands, and the protein can be used for inhibiting activation of hemolymph PPO of pieris rapae or papilio xuthuslinnaeus.

Description

technical field [0001] The invention relates to the fields of molecular biology, genetic engineering and protein engineering. In particular, a protein PpVPG expressed in the chrysalis golden wasp venom and its encoded nucleic acid sequence and application. Background technique [0002] Human beings have been engaged in agricultural activities for tens of thousands of years. The domestication of plants has caused the problem of pest control. People have been looking for effective methods to control pests for a long time. Through years of exploration, integrated pest management strategies have been proposed in order to achieve economic harm Under the allowable level, the best economic, ecological and social benefits can be obtained. At present, chemical control is mostly used in pest control. From the artificial synthesis of DDT in 1939 to organic phosphorus and other pesticides and carbamates, chemical pesticides became the main means of early pest control. However, the lon...

Claims

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Application Information

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IPC IPC(8): C07K14/435C12N15/12
CPCC07K14/43572C07K2319/02
Inventor 叶恭银王蓓蓓方琦刘明明
Owner ZHEJIANG UNIV
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