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Insecticidal protein, and coding gene and use thereof

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, environmental pollution, increasing production costs, etc.

Active Publication Date: 2014-07-09
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 virulence of PIC3 insecticidal protein in plants

Method used

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

Examples

Experimental program
Comparison scheme
Effect test

no. 1 example

[0078] The first embodiment, the acquisition and synthesis of PIC3-01 gene sequence

[0079] 1. Obtain the PIC3-01 gene sequence

[0080] The amino acid sequence (731 amino acids) of the PIC3-01 insecticidal protein, as shown in SEQ ID NO:2 in the sequence listing; obtain the amino acid sequence (731 amino acids) corresponding to the coding of the PIC3-01 insecticidal protein according to the maize preference codon amino acids) nucleotide sequence (2196 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.

[0081] 2. Synthesize the above-mentioned PIC3-01 nucleotide sequence

[0082] The PIC3-01 nucleotide sequence (shown as SEQ ID NO: 1 in the sequence listing) was synthesized by Nanjing GenScript Biotechnology Co., Ltd.; the synthesized PIC3-01 nucleotide sequence (SEQ ID NO: 1 ) is also connected with a SphI restriction site, and the 3' end of the PIC...

no. 2 example

[0086] The second embodiment, construction of recombinant expression vector and transformation of recombinant expression vector into Agrobacterium

[0087] 1. Construction of recombinant cloning vector DBN01-T containing PIC3-01 nucleotide sequence

[0088] The synthetic PIC3-01 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 initiation codon; SP6 is the SP6 RNA polymerase promoter; T7 is the T7 RNA polymerase promoter; PIC3-01 is the PIC3 -01 nucleotide sequence (SEQ ID NO: 1); MCS is a multiple cloning site).

[0089]Then, the recombinant cloning vector DBN01-T was transformed in...

no. 3 example

[0105] The third embodiment, the acquisition and verification of maize plants transferred to the PIC3-01 nucleotide sequence

[0106] 1. Obtaining corn plants transferred to the PIC3-01 nucleotide sequence

[0107]According to the routinely adopted Agrobacterium infection method, the immature embryos of the aseptically cultured corn variety Zong 31 (Z31) were co-cultured with the Agrobacterium described in 4 in the second embodiment, so that 2 and 3 in the second embodiment T-DNA (including the promoter sequence of corn Ubiquitin gene, PIC3-01 nucleotide sequence, PIC3-01 Substitution nucleotide sequence, PIC3-01 truncated nucleotide sequence, PIC3-01 added nucleotide sequence, natural sequence, PMI gene and Nos terminator sequence) were transferred into the maize genome, and the transferred PIC3- Corn plant with 01 nucleotide sequence, corn plant with PIC3-01 substituted nucleotide sequence, corn plant with PIC3-01 truncated nucleotide sequence, corn plant with PIC3-01 added...

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Abstract

The invention relates to an insecticidal protein, and a coding gene and use thereof. The insecticide protein comprises (a) a protein formed by an amino acid sequence shown as SEQ ID NO:2; or (b) a protein derived from (a), wherein the protein is substituted and / or deleted and / or added with one or more amino acids in the amino acid sequence in (a) and has the insecticidal activity; or (c) a protein formed by the amino acid sequence which has at least 90% of sequence homology with the SEQ ID NO:2. The insecticide protein provided by the invention is not only high in expression quantity, but also good in stability, and is strong in toxicity to injurious insects.

Description

technical field [0001] The present invention relates to an insecticidal protein, its encoding gene and application, in particular to a modified PIC3-01 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
Inventor 张爱红
Owner BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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