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Common mussel C type lysozyme and preparation method thereof

A technology of blue mussel and lysozyme, which is applied in the field of lysozyme, can solve the problem that there is no antibacterial activity of blue mussel C-type lysozyme, and achieve the effect of strong inhibitory activity

Inactive Publication Date: 2012-08-22
NINGBO UNIV
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Problems solved by technology

So far, there is no research report on C-type lysozyme and its antibacterial activity

Method used

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Embodiment

[0018] 1. Use Trizol reagent to extract total RNA from purple mussel hemolymph infected by Vibrio anguillarum. The extraction method is carried out according to the instructions of Invitrogen’s Trizol reagent: take 50 ml of purple mussel hemolymph, centrifuge at 2000 g for 10 minutes, discard Add 20 ml of TRIZOL (Invitrogen) to the precipitated cells, disperse the cells with a high-speed disperser, shake vigorously at room temperature for 10 seconds; add 4 ml chloroform, shake vigorously for 30 seconds; centrifuge at 10,000 g at 4°C for 10 minutes; draw Put the supernatant into a new centrifuge tube, add an equal volume of isopropanol (approximately 15ml) that has been ice-bathed, and place it at -20°C for more than 1 hour; centrifuge at 10,000 g at a high speed for 10 minutes at 4°C; carefully discard the supernatant Wash the precipitate with 5 ml of 70% ethanol; centrifuge at 10,000g at 4°C for 10 minutes at high speed, discard the supernatant carefully; dry in vacuum for 5 m...

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Abstract

The invention discloses a common mussel C type lysozyme and a preparation method thereof. The amino acid sequence of the common mussel C type lysozyme is shown by a sequence table SEQ ID No.1. The preparation method of the common mussel C type lysozyme comprises the following steps of: extracting common mussel gross RNA (ribonucleic acid), purifying mRNA (Messenger ribonucleic acid), preparing a cDNA (complementary deoxyribonucleic acid) template solution, cloning gene, building recombinant plasmid, expressing expression plasmid and purifying protein to obtain the C type lysozyme. The common mussel C type lysozyme is a new C type lysozyme which has strong inhibitory activity on various gram positive and negative microorganisms, and has application value in the aspects of developing food preservatives, feed additive, etc.

Description

technical field [0001] The invention relates to lysozyme, in particular to C-type lysozyme from purple mussel and a preparation method thereof. Background technique [0002] Lysozyme (Lysozyme, EC 3.2.1.17) is one of the most important innate immune factors in invertebrates. It mainly cuts off the β-1 between N-acetylglucosamine and N-acetylmuramic acid in peptidoglycan , The link between the 4 glycosidic bonds destroys the peptidoglycan scaffold, causing the cells to burst and cause bacterial lysis. Lysozyme can not only directly kill bacteria during the body's immune response, but also induce and regulate the synthesis and secretion of other immune factors. Lysozyme can be divided into six types according to its source: C (chicken) type, G (goose) type, I (invertebrate) type, plant type, bacterial type and T4 phage type lysozyme. G-type lysozyme mainly exists in mammals, birds and fish, and the patent numbers are CN 2004100506518 and 2004100506522. G-type lysozyme was ob...

Claims

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Application Information

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IPC IPC(8): C12N9/36C12N15/56C12N15/70C12R1/19
Inventor 王春艳母昌考宋微微李荣华王春琳
Owner NINGBO UNIV
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