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Chitinase capable of degrading spore walls of myxosporean and encoding genes thereof

A chitinase, coding gene technology, applied in genetic engineering, plant genetic improvement, enzymes, etc., can solve problems such as myxosporidium that have not yet been seen

Active Publication Date: 2009-12-16
FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, as the main biological enzyme of chitin degradation, chitinase has been studied a lot, and a complete enzymatic research method has been established, but it has not been applied to the degradation of chitin shell valves of Myxosporum report

Method used

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  • Chitinase capable of degrading spore walls of myxosporean and encoding genes thereof
  • Chitinase capable of degrading spore walls of myxosporean and encoding genes thereof
  • Chitinase capable of degrading spore walls of myxosporean and encoding genes thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1. Acquisition of chitinase and its coding gene for myxospore spore wall degradation ability

[0040] 1. Screening of chitinase-producing strains and cloning of chitinase gene fragments

[0041] 1. Experimental materials

[0042] 1) Material

[0043] DNA Molecular Weight Marker was purchased from Tiangen Biochemical Technology (Beijing) Co., Ltd.

[0044] The chitinase substrate chitin is a product of Sigma Company. Yeast extract and peptone were purchased from Oxford Company. The rest of the chemical reagents were of domestic analytical grade.

[0045] 2) solution

[0046] (1) PBS buffer: NaCl 137mmol / L, KCl 2.7mmol / L, NaCl 2 HPO 4 4.3mmol / L, KH 2 PO 4 1.4mmol / L, pH 7.2.

[0047] (2) 50×TAE: 242g Tris base, 57.1mL glacial acetic acid, 100mL EDTA (0.5mol / L, pH 8.0), dissolved in deionized water and adjusted to 1000mL.

[0048](3) Lysozyme mixture: 2 mg / mL lysozyme, 50 μg / mL RNaseA, 0.3 mol / L sucrose, 25 mmol / L Tris-HCl (pH 8.0), 25 mmol / L EDTA (pH 8...

Embodiment 2

[0170] Example 2. Functional verification of chitinase of the present invention and its coding gene's ability to degrade myxospore spore wall

[0171] 1. Expression of chitinase gene ChiCD3 in Escherichia coli

[0172] 1 Experimental materials

[0173] 1) Strains, vectors, tool enzymes, reagents

[0174] Escherichia coli BL21 (DE3) competent cells were purchased from Tiangen Biochemical Technology (Beijing) Co., Ltd. The pET-30a(+) vector is the product of Novagen.

[0175] DNA agarose gel recovery kit and DNA purification kit are products of TakaRa company, Taq DNA polymerase used in the experiment was purchased from TakaRa company, various restriction endonucleases and T 4 DNA ligase was purchased from NEB Company, and DNA molecular weight standards were purchased from Tiangen Biochemical Technology (Beijing) Co., Ltd.

[0176] The substrate chitin is the product of Sigma Company, the protein low molecular weight standard is purchased from Shanghai Institute of Biochemi...

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Abstract

The invention discloses chitinase capable of degrading spore walls of myxosporean and encoding genes thereof. The chitinase is a protein with either of the following amino acid residue sequence: 1) SEQ ID No. 2 the amino acid residue sequence in a sequence table; and 2) sequence of a protein which has the related function of the chitinase and is derived from SEQ ID No. 2 by replacing and / or deleting and / or adding one or more amino acid residues in the amino acid residue sequence. Experiments show that the chitinase has the activity for degrading chitin, and can destroy the spore walls of the myxosporean. Therefore, the chitinase and the encoding genes thereof can be used for preventing and controlling myxosporidiosis, particularly fish myxosporidiosis.

Description

technical field [0001] The invention relates to a chitinase capable of degrading myxospore spore wall and its coding gene. Background technique [0002] Myxosporidia (Myxosporidia), the general name of Myxosporidia, is a common protozoan parasite in ectothermic animals. There are many types. There are nearly a thousand species reported now, except for a very small number of parasites in amphibians and reptiles. , the vast majority of which are obligate fish parasites, parasitic in fish internal and external tissues or lacunar organs. The myxosporidosis caused by the parasitism of myxospores has caused huge economic losses to the aquaculture industry (Zhang Jinyong, Wu Yingsong, Lu Yishan, Wang Jianguo. Research progress of fish microsporidia. Acta Hydrobiology. 2004, 5 :545-568). [0003] There are many kinds of myxospores, and new species are discovered almost every year, each of which will have different degrees of impact on its host fish, so myxosporidosis is extremely ...

Claims

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Application Information

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IPC IPC(8): C12N9/42C12N15/56C12N15/63C12N5/10C12N1/00A01N63/02A01P1/00C12N1/20C12R1/01
Inventor 周志刚姚斌刘玉春何夙旭曹雅男白东清石鹏君孟昆
Owner FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES
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