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Antimicrobial peptide as well as preparation method and application thereof

An antibacterial peptide and signal peptide technology, applied in the field of anti-nematode peptide and its preparation, can solve the problems of limited resources, easy loss of resistance, easy to be affected by the environment, etc.

Inactive Publication Date: 2013-12-11
INST OF VEGETABLE & FLOWERS CHINESE ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Biological control has a wide potential of biological resources, and has certain safety for the environment and people, but the effect of biological control is unstable and easily affected by the environment, and it is not easy to be commercialized. At present, only a few strains are used for biological control
The use of anti-nematode genes is an efficient measure to control plant-parasitic nematodes, but the number of anti-nematode genes found now is very small, resources are limited, and there are problems such as the differentiation of nematode virulence populations and the easy loss of resistance.

Method used

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  • Antimicrobial peptide as well as preparation method and application thereof
  • Antimicrobial peptide as well as preparation method and application thereof
  • Antimicrobial peptide as well as preparation method and application thereof

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preparation example Construction

[0043] The method for preparing the antimicrobial peptide developed by the present invention comprises the following steps, (1) adopting the trizol method to extract the total RNA of the pepper leaf tissue, and adopting reverse transcriptase to obtain the first strand cDNA for subsequent use; the reverse transcription used Enzyme: First-Strand Synthesis Kit (Invitrogen).

[0044] (2) According to the sequence of the known antimicrobial peptide Snakin gene, design upstream primers and downstream primers, use the first-strand cDNA as a template, clone the gene coding sequence without signal peptide in the antimicrobial peptide Snakin through PCR technology, and perform sequencing to obtain the complete The long gene sequence is recorded as the Sn gene sequence; the upstream primer sequence is 5'ATT CAA ACT GAT CAT GTT GC3', and the downstream primer sequence is 5'TTA AGG GCA TTT GCG TCT3';

[0045] (3) Cloning the Sn gene sequence obtained in step (2) into the prokaryotic expres...

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Abstract

The invention discloses an antimicrobial peptide. The gene sequence of the antimicrobial peptide is a gene code sequence of a no-signal peptide of an antimicrobial peptide Snakin. The invention further discloses a method for preparing the antimicrobial peptide. The method comprises the following steps of (1) extracting the total RNA (Ribonucleic Acid) of capsicum leaf tissue, thereby obtaining a first chain cDNA (complementary Deoxyribonucleic Acid) for standby by adopting a reverse transcriptase; (2) carrying out PCR (Polymerase Chain Reaction) cloning to obtain the gene code sequence, denoted by an Sn gene sequence, of the no-signal peptide of the antimicrobial peptide Snakin; (3) forming recombination plasmids; (4) screening successfully-recombined plasmids denoted by pET-Sn; (5) transforming the successfully-recombined plasmids into competent cells BL21 (DE3) of Escherichia coli, carrying out induced expression by adopting IPTG, and meanwhile, secreting proteins; (6) carrying out ultrasonic-wave crushing on the competent cells which secrete the proteins in the step (5), carrying out centrifugation, and then, collecting the antimicrobial peptide from an supernatant. The method has the advantages that the antimicrobial peptide which does not contain signal peptides is obtained and has a killing effect on meloidogyne incognita chitwood.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to an anti-nematode peptide and its preparation method and application. Background technique [0002] Plant parasitic nematode is an important disease in agricultural production. It is a worldwide disease. It is highly destructive, has many types of hosts, and has a wide range of distribution. It is extremely difficult to control and seriously hinders the sustainable development of agriculture. It causes 157 billion Dollar economic loss. [0003] At present, the control of plant parasitic nematodes usually takes measures such as chemical nematicides, cultivation techniques and breeding resistant varieties. However, due to the food safety and environmental problems of chemical nematicides, their use is strictly controlled. The host range of root-knot nematode can reach more than 3,000 species, and it mainly harms the roots of plants. Therefore, the control of root-knot nematode through...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/70A01N47/44A01P5/00
Inventor 茆振川张文玥凌键陈国华杨宇红谢丙炎
Owner INST OF VEGETABLE & FLOWERS CHINESE ACAD OF AGRI SCI
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