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Insecticidal protein, coding gene of insecticidal protein and purpose of insecticidal protein

A technology of insecticidal protein and insecticidal gene, applied in the field of insecticidal protein, can solve problems affecting the living conditions of the population, undiscovered PIC9 insecticidal protein expression level and virulence, environmental pollution, etc.

Active Publication Date: 2012-11-21
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 PIC9 insecticidal protein in plants

Method used

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  • Insecticidal protein, coding gene of insecticidal protein and purpose of insecticidal protein
  • Insecticidal protein, coding gene of insecticidal protein and purpose of insecticidal protein
  • Insecticidal protein, coding gene of insecticidal protein and purpose of insecticidal protein

Examples

Experimental program
Comparison scheme
Effect test

no. 1 example

[0073] The first embodiment, the acquisition and synthesis of PIC9-02 gene sequence

[0074] 1. Obtain the PIC9-02 gene sequence

[0075] The amino acid sequence (699 amino acids) of the PIC9-02 insecticidal protein is shown in SEQ ID NO: 2 in the sequence listing; the amino acid sequence (699 amino acids) encoding the insecticidal protein corresponding to the PIC9-02 is obtained according to the maize preference codon amino acids) nucleotide sequence (2100 nucleotides), as shown in SEQ ID NO:1 in the sequence listing. For maize codon usage bias, please refer to http: / / www.kazusa.or.jp / codon / cgi-bin / showcodon.cgi?species=381124.

[0076] 2. Synthesize the above-mentioned PIC9-02 nucleotide sequence

[0077] The PIC9-02 nucleotide sequence (shown as SEQ ID NO: 1 in the sequence listing) was synthesized by Nanjing GenScript Biotechnology Company; the synthesized PIC9-02 nucleotide sequence (SEQ ID NO: 1) The 5' end of the PIC9-02 nucleotide sequence (SEQ ID NO: 1) is also con...

no. 2 example

[0081] The second embodiment, construction of recombinant expression vector and transformation of recombinant expression vector into Agrobacterium

[0082] 1. Construction of recombinant cloning vector DBN01-T containing PIC9-02 nucleotide sequence

[0083] The synthesized PIC9-02 nucleotide sequence was connected to 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 SP6 RNA polymerase promoter; T7 is the T7 RNA polymerase promoter; PIC9-02 is the PIC9-02 nucleotide sequence (SEQ ID NO: 1); MCS is the multiple cloning site).

[0084] Then, the recombinant cloning vector DBN01-T was transformed int...

no. 3 example

[0100] The third embodiment, the acquisition and verification of corn plants transferred to PIC9-02 nucleotide sequence

[0101] 1. Obtain corn plants transferred to PIC9-02 nucleotide sequence

[0102] According to the routinely used Agrobacterium infection method, the immature embryos of the aseptically cultivated corn variety Zong 31 (Z31) were co-cultured with the Agrobacterium described in 4 in the second example, so as to infect 2 and 3 in the second example. T-DNA (including the promoter sequence of corn Ubiquitin gene, PIC9-02 nucleotide sequence, PIC9- 02 substituted nucleotide sequence, PIC9-02 deleted nucleotide sequence, PIC9-02 added nucleotide sequence, known sequence, PMI gene and Nos terminator sequence) were transferred into the maize genome, and the transferred PIC9 -02 nucleotide sequence corn plant, corn plant transferred to PIC9-02 substituted nucleotide sequence, corn plant transferred to PIC9-02 deleted nucleotide sequence, corn plant transferred to PIC...

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PUM

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Abstract

The invention relates to insecticidal protein, a coding gene of the insecticidal protein and a purpose of the insecticidal protein. The insecticidal protein comprises (a) protein consisting of amino acid sequences shown by SEQ ID NO. 2; or (b) protein which has the insecticidal activity and is derived from (a) through substituting, deleting or adding one or several amino acids from / into the amino acid sequences of (a); or (c) protein generated through expression of the nucleic acid molecule containing the nucleotide sequences of SEQ ID NO. 1; or (d) protein generated through expression of the nucleic acid molecule containing nucleotide sequences, wherein the nucleotide sequences have complementary sequences hybridized with the nucleotide sequences of SEQ ID NO. 1 under the strict conditions; or (e) protein generated through expression of the nucleic acid molecule containing nucleotide sequences, wherein the nucleotide sequences have the similar codes to the nucleotide sequences of (d). The insecticidal protein has high expression quantity and strong toxicity on insect pests.

Description

technical field [0001] The present invention relates to an insecticidal protein, its encoding gene and application, in particular to a modified PIC9-02 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 Applications(China)
IPC IPC(8): C07K14/325C12N15/32C12N15/63C12N1/13C12N1/19C12N1/21C12N5/10C12N7/01C12P21/02A01K67/033A01H5/00A01N63/02A01P7/04A01N63/50
CPCC12N15/8286A01N63/00A01N63/02A01N37/44C07K14/325A01N37/46Y02A40/146A01N63/50
Inventor 张爱红庞洁牛瑞琪牛丽红王娜黄金存祁玉洁
Owner BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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