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Method for mutagenesis breeding neuter protease high yield bacterial strain by hypophrenia N+ion injection technology

A neutral protease, ion implantation technology, applied in the fields of genetic engineering and enzymology, can solve the problem of high-yielding strains of neutral protease, etc., and achieve the effects of good economic benefits, wide mutation spectrum and high mutation rate

Inactive Publication Date: 2008-05-21
TIANJIN UNIV OF SCI & TECH
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Problems solved by technology

[0004] At present, no method has been found to use ion implantation technology to mutagenize and breed strains with high production of neutral protease

Method used

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  • Method for mutagenesis breeding neuter protease high yield bacterial strain by hypophrenia N+ion injection technology

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Embodiment Construction

[0021] The present invention will be further described by the following examples; the following examples are illustrative, not limiting, and cannot limit the protection scope of the present invention with the following examples.

[0022] A low-energy N + The method for ion implantation technology mutagenesis selection and breeding of high-yield strains of neutral protease, the steps are:

[0023] 1. Screening of starting strains

[0024] Pick a ring of Bacillus subtilis AS1.398 from the slant and insert it into the seed medium. After culturing at 32°C and 180r / min for 24h, take 0.1mL and spread it on a plate, and continue culturing at 32°C for 48h. Then pick 20 single colonies from the plate and transfer them to the slant, put these 20 strains into the seed culture solution and cultivate them for 24 hours, measure the activity of neutral protease, and select the strain with higher and more stable enzyme activity as the starting strain. The results are (see Table 1): after th...

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Abstract

The invention relates to an inducing and breeding method of high yield neutral proteinase strain by using low energy N <+> ion injection technology. The steps are that (1) the initial strain is screened; (2) the N <+> ion is injected for the induced mutation; (3) the high yield strain is screened; (4) the N <+> ion is injected for the induced mutation; (5) the high yield neutral proteinase strain is determined. The invention uses the N <+> ion injection technology for the induced mutation of Bacillus Subtilis AS1.398 of neutral proteinase; after the injection of different dosage of N <+> ion, the survival rate of the strain takes a typical saddle shape dose-effect curve; a high yield mutant strain with good genetic stability can be screened at the best injection dosage of 50*10 <14> ions / cm <2>; the shaking flask activity of the neutral proteinase is about 8230U / mL, which is improved by 81.3 percent. The ion injection technology can be applied into the mutation and selection of high yield neutral proteinase strain; the invention has a higher mutation rate and a wider mutation spectrum for the microorganism; the invention has good mutation effects, which is an ideal breeding method for the microorganism.

Description

technical field [0001] The present invention relates to the field of genetic engineering and enzymology, more specifically, relates to a method using low-energy N + A method for the mutagenesis and breeding of high-producing strains of neutral protease by ion implantation technology. Background technique [0002] Neutral protease refers to a class of protease with the optimum pH between 6.0 and 7.5, which can catalyze the hydrolysis of protein peptide bonds. Because of its fast catalytic reaction speed, no industrial pollution, wide adaptability of catalytic reaction conditions and other properties and advantages, it is widely used in food, pharmaceutical, cosmetics, detergent, silk spinning, fur softening and other industries. Neutral proteases can be extracted from animal and plant tissues, but due to the limitations of this method, people are increasingly focusing on microbial proteases. [0003] In recent years, ion beam, as a new mutagenic source, has developed extrem...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/01C12N13/00C12N1/20C12R1/125
Inventor 路福平杜连祥张敏王海宽李玉王春霞王建玲吴雅倩
Owner TIANJIN UNIV OF SCI & TECH
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