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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 the problems of repeated application, increased production cost, and impact on the living conditions of the population.

Active Publication Date: 2014-04-23
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] First Example, 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) corresponding to the PIC10-01 insecticidal protein was obtained according to the rice-preferred codons. amino acid) nucleotide sequence (1896 nucleotides), as shown in SEQ ID NO: 1 in the sequence listing. The PIC10-01 nucleotide sequence (as shown in 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 is also connected with a SpeI restriction site, and the 3' end of the PIC10-01 nucleotide sequence (SEQ ID NO: 1) is also connected with a PvuI restriction site.

no. 2 example

[0075] 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 , the construction process is as follows figure 1 (wherein, Amp is the ampicillin resistance gene; f1 is the replication origin of phage f1; LacZ is the LacZ initiation codon; SP6 is the SP6 RNA polymerase promoter; T7 is the T7 RNA polymerase promoter; PIC10-01 is the PIC10 -01 nucleotide sequence (SEQ ID NO: 1); MCS is a multiple cloning site).

[0078] Then the recombinant cloning vector DBN01-T was transformed into E. coli T1 compet...

no. 3 example

[0088] The third example, the acquisition and verification of rice plants transformed with PIC10-01 nucleotide sequence

[0089] 1. Obtain rice plants transformed with PIC10-01 nucleotide sequence

[0090] According to the conventional 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 recombinants constructed in 2 and 3 in the second embodiment were co-cultured. 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 the rice chromosome In the group, rice plants transformed with the PIC10-01 nucleotide sequence and rice plants with known sequences were obtained; at the same time, wild-type rice plants were used as controls.

[0091] For Agrobacterium-medi...

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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 crop losses, 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 pesticides and biological pesticides, but both have limitations in practical application: chemical pesticides will bring environmental pollution problems and lead to the development of drug-resistant insects. However, bio-insecticides are easily degraded in the environment and require repeated application in production, which greatly increases production costs. [0003] In order to solve the limitations of chemical pesticides and biological pesticides in practical applications, scientists have foun...

Claims

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Application Information

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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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