Insecticidal protein, and encoding gene and use thereof

A technology of insecticidal protein and insecticidal gene, applied in the field of insecticidal protein, can solve problems such as environmental pollution, affecting the living conditions of the population, and farmers' economic losses.

Active Publication Date: 2015-06-17
BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002]Pest pests are the main factor leading to crop loss, causing significant economic losses to farmers, and even affecting the survival of the local population
In order to control plant pests, people usually use broad-spectrum chemical insecticides and biological insecticides, but both have limitations in practical application: chemical insecticides will cause environmental pollution problems and lead to the emergence of resistant insects. However, biopesticides are easily degraded in the environment and need to be repeatedly applied in production, which greatly increases the production cost
There is no report on the expression level and toxicity of PIC10 insecticidal protein in plants

Method used

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  • Insecticidal protein, and encoding gene and use thereof
  • Insecticidal protein, and encoding gene and use thereof
  • Insecticidal protein, and encoding gene and use thereof

Examples

Experimental program
Comparison scheme
Effect test

no. 1 example

[0073] The first embodiment, the acquisition and synthesis of PIC10-01 gene sequence

[0074] The amino acid sequence (630 amino acids) of the PIC10-01 insecticidal protein is shown in SEQ ID NO: 2 in the sequence listing; the amino acid sequence (630 amino acids) encoding the insecticidal protein corresponding to the PIC10-01 is obtained according to rice preference codons amino acid) nucleotide sequence (1896 nucleotides), as shown in SEQID NO:1 in the sequence listing. The PIC10-01 nucleotide sequence (shown as SEQ ID NO: 1 in the sequence listing) was synthesized by Nanjing GenScript Biotechnology Co., Ltd.; the synthesized PIC10-01 nucleotide sequence (SEQ ID NO: 1) The 5' end of the PIC10-01 nucleotide sequence (SEQ ID NO: 1) is also connected with a PvuI restriction site at the 3' end.

no. 2 example

[0075] The second embodiment, construction of recombinant expression vector and transformation of recombinant expression vector into Agrobacterium

[0076] 1. Construction of recombinant cloning vector DBN01-T containing PIC10-01 nucleotide sequence

[0077] The synthesized PIC10-01 nucleotide sequence was ligated into the cloning vector pGEM-T (Promega, Madison, USA, CAT: A3600), and the operation steps were carried out according to the instructions of the pGEM-T vector produced by Promega Company to obtain the recombinant cloning vector DBN01-T , its construction process is as follows figure 1 Shown (wherein, Amp represents the ampicillin resistance gene; f1 represents the replication origin of phage f1; LacZ is the LacZ start codon; SP6 is the promoter of SP6 RNA polymerase; T7 is the promoter of T7 RNA polymerase; PIC10-01 is the promoter of PIC10 -01 nucleotide sequence (SEQ ID NO: 1); MCS is a multiple cloning site).

[0078] Then, the recombinant cloning vector DBN01-...

no. 3 example

[0088] The third embodiment, the acquisition and verification of rice plants transferred to the PIC10-01 nucleotide sequence

[0089] 1. Obtain rice plants transferred to the PIC10-01 nucleotide sequence

[0090] According to the routinely used Agrobacterium infection method, the callus of the aseptically cultured japonica rice variety Nipponbare was co-cultured with the Agrobacterium described in 4 in the second embodiment, so that the recombinant strains constructed in 2 and 3 in the second embodiment The T-DNA (including the promoter sequence of maize Ubiquitin gene, PIC10-01 nucleotide sequence, known sequence, PMI gene and Nos terminator sequence) in the expression vectors DBN100323 and DBN100323N (known sequence) was transferred into rice chromosome In the group, the rice plants transformed with the PIC10-01 nucleotide sequence and the rice plants transformed with the known sequence were obtained; at the same time, wild-type rice plants were used as controls.

[0091] F...

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PUM

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Abstract

The invention relates to an insecticidal protein, and an encoding gene and use thereof. The insecticidal protein comprises (a) protein formed by an amino acid sequence shown in SEQ ID NO:2, or (b) protein which has insecticidal activity and is derived from (a) in a manner of substituting and / or deleting and / or adding one or more amino acids to the amino acid sequence in (a). The insecticidal protein disclosed by the invention is especially applicable to expression in monocotyledons, especially rice, so that not only are the expression quantity and the stability of PIC10-01 insecticidal protein significantly improved, but also the toxicity of the PIC10-01 insecticidal protein on insects and injurious insects, especially lepidoptera insect pests is significantly enhanced.

Description

technical field [0001] The present invention relates to an insecticidal protein, its encoding gene and application, in particular to a modified PIC10-01 insecticidal protein, its encoding gene and application. Background technique [0002] Plant pests are the main factor leading to the loss of crops, causing significant economic losses to farmers, and even affecting the living conditions of the local population. In order to control plant pests, people usually use broad-spectrum chemical insecticides and biological insecticides, but both have limitations in practical application: chemical insecticides will cause environmental pollution problems and lead to the emergence of resistant insects. However, biopesticides are easily degraded in the environment and require repeated application in production, which greatly increases production costs. [0003] In order to solve the limitations in the practical application of chemical insecticides and biological insecticides, scientists...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/325C12N15/32C12N15/63C12N5/10C12N1/13C12N1/19C12N1/21C12N7/01C12P21/02A01K67/033A01H5/00A01N47/44A01P7/04
CPCA01N47/44C07K14/325C12N15/8286
Inventor 韩超庞洁丁德荣李胜兵
Owner BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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