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Insecticide gene and application thereof

A technology for killing insects and pests, which can be used in the fields of insecticides, genetic engineering, plant genetic improvement, etc., and can solve the problems of unsatisfactory expression and virulence

Active Publication Date: 2014-09-17
BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to provide a kind of insecticidal gene and its application, which is optimized and transformed according to the preferred codon of corn, and effectively overcomes the unsatisfactory expression and toxicity of the Cry1Ab insecticidal protein in corn in the prior art and other technical defects

Method used

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  • Insecticide gene and application thereof
  • Insecticide gene and application thereof
  • Insecticide gene and application thereof

Examples

Experimental program
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Effect test

no. 1 example

[0046] First embodiment, selection, optimization and synthesis of natural Cry1Ab gene sequence

[0047] 1. Select the natural Cry1Ab gene sequence for optimization

[0048] The nucleotide sequence of the natural Cry1Ab8 gene with the serial number AAA22551 registered by Oeda et al. in 1993 was selected as the basis for the optimization and modification of the insecticidal gene of the present invention. The nucleotide sequence of the natural Cry1Ab8 gene is as SEQ ID NO: 2 in the sequence table. As shown, the amino acid sequence of the native Cry1Ab8 gene is shown in SEQ ID NO: 3 in the sequence listing.

[0049] 2. Intercept the natural Cry1Ab gene sequence for optimization and transformation

[0050] The nucleotide sequence corresponding to positions 1 to 615 of the amino acid sequence encoded by the nucleotide sequence of the above-mentioned natural Cry1Ab8 gene was cut out.

[0051] 3. Optimize the above-mentioned truncated natural Cry1Ab gene sequence

[0052] Keeping t...

no. 2 example

[0057] Second embodiment, construction of recombinant expression vector and transformation of recombinant expression vector into Agrobacterium

[0058] 1. Construction of the recombinant cloning vector DBN01Ab-T containing the optimized Cry1Ab gene sequence

[0059] The synthesized optimized Cry1Ab gene 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 product of Promega Company to obtain the recombinant cloning vector DBN01Ab-T. Its construction Process such as figure 1(wherein, Amp represents ampicillin resistance gene; f1 represents the origin of replication of phage f1; LacZ is the LacZ initiation codon; SP6 is the SP6 RNA polymerase promoter; T7 is the T7 RNA polymerase promoter; mCry1Ab is the optimized Cry1Ab gene sequence (SEQ ID NO: 1); MCS is the multiple cloning site).

[0060] Then the recombinant cloning vector DBN01Ab-T was transfor...

no. 3 example

[0070] The third embodiment, the acquisition and verification of the maize plant transformed into the optimized Cry1Ab gene sequence

[0071] 1. Obtain the maize plant transformed into the optimized Cry1Ab gene sequence

[0072] According to the conventional Agrobacterium infection method, the aseptically cultured immature embryos of maize variety Zong 31 (Z31) were co-cultivated with the Agrobacterium described in 4 in the second example, so that the 2 and 3 in the second example were co-cultured. The constructed recombinant expression vectors DBN100053 and DBN100053R (known sequences) T-DNA (including the promoter sequence of the maize Ubiquitin gene, the optimized Cry1Ab gene sequence, the known optimized Cry1Ab gene sequence, the PMI gene and the Nos terminator sequence) were transfected. Into the maize genome, a maize plant transformed with the optimized Cry1Ab gene sequence and a maize plant with a known optimized Cry1Ab gene sequence (positive control) were obtained; at...

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Abstract

The invention relates to insecticide gene and application thereof. A nucleotide sequence of the insecticide gene comprises a sequence (a) or a sequence (b), wherein the (a) has a nucleotide sequence shown in an SEQ ID NO: 1, and the (b) is a complementary nucleotide sequence with the nucleotide sequence limited by the (a). The insecticide gene is suitable for being expressed in corns, not only remarkably improves protein expression index and stability of CrylAb insecticide protein, and remarkably enhances toxicity of the CrylAb insecticide protein on main insects of corns.

Description

technical field [0001] The present invention relates to an insecticidal gene and use thereof, in particular to a modified Cry1Ab insecticidal gene and use thereof. 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 found that some insect-resistant transgenic plants...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/32C12N15/63C12N5/10C12N1/21C12N1/19C12N7/01C12N15/82A01H5/00A01N47/44A01P7/04
Inventor 丁德荣庞洁康越景杨进孝贾志伟黄金存郭函子
Owner BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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