Method for breeding bacillus natto and lactobacillus casei shirota to ferment LactucapsativapL.var.asparaginapBailey

A technology of Bacillus natto and Lactobacillus casei, applied in the direction of Lactobacillus, microorganism-based methods, biochemical equipment and methods, etc., can solve the problem of unrelated breeding medium and fermentation medium, low yield of target products, etc. question

Inactive Publication Date: 2015-03-11
HANGZHOU YUANPEITE BIOTECH
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0025] However, there are certain defects in the strategy of genome rearrangement at present, mainly because there is no connection between the breeding medium and the fermentation medium, so that the yield of the target product of the selected strains during fermentation is much lower than expected

Method used

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  • Method for breeding bacillus natto and lactobacillus casei shirota to ferment LactucapsativapL.var.asparaginapBailey
  • Method for breeding bacillus natto and lactobacillus casei shirota to ferment LactucapsativapL.var.asparaginapBailey
  • Method for breeding bacillus natto and lactobacillus casei shirota to ferment LactucapsativapL.var.asparaginapBailey

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preparation example Construction

[0102] (h) The lettuce juice is prepared by directly squeezing the lettuce juice.

[0103] In the embodiment of the present invention, Bacillus natto 10261 and Bacillus natto 10263 adopt the prior art, and Bacillus natto 10261 comes from China Industrial Microorganism Culture Collection Center (CICC), address: 24 Jiuxianqiao Middle Road, Chaoyang District, Beijing No. 6 building; the storage date is 2002, and the strain maintenance number is 10261; Bacillus natto 10263 is from the China Industrial Microbiology Culture Collection Center (CICC), address: Courtyard 6, No. 24, Jiuxianqiao Middle Road, Chaoyang District, Beijing Building No. 1; the storage date was 2002, and the strain maintenance number was 10263; Lactobacillus casei Shirota was from Yakult (Shanghai) Co., Ltd.; the mouse anti-PQQ IgG monoclonal antibody PQQ-mAb was a commercially available product, purchased from Jiaxing Yuanke Biotechnology Co., Ltd., the product number is YK-Ab-013.

Embodiment 1

[0105] The present invention prepares fermented lettuce rich in NK and PQQ through the breeding of Bacillus natto and Lactobacillus casei and their co-fermentation, comprising the following steps:

[0106] (1) Breeding of Bacillus natto BN-NKPQQ-RWX-406 with high yield of NK and PQQ

[0107] Using the substrate-induced adaptive strategy, the high-yield NK and PQQ Bacillus natto BN-NKPQQ-RWX-406 was obtained through genome rearrangement and high-throughput screening technology. The obtained Bacillus natto 10261 and 10263 producing NK and PQQ were subjected to nitrosoguanidine mutagenesis respectively; then the offspring were mixed for mutagenesis, and cell fusion was carried out (fusogenic agent was polyethylene glycol 4000), subcultured, Realized genome shuffling, optimized and integrated the dominant mutations of all mutagenized progenies of Bacillus natto 10261 and 10623, and bred a high-yield NK and PQQ Natto with high yield, vigorous growth, stable genetics, and suitable a...

Embodiment 2

[0119] The present invention prepares fermented lettuce rich in NK and PQQ through the breeding of Bacillus natto and Lactobacillus casei and their co-fermentation, comprising the following steps:

[0120] (1) Breeding of Bacillus natto BN-NKPQQ-RWX-406 with high yield of NK and PQQ

[0121] Using the substrate-induced adaptive strategy, the high-yield NK and PQQ Bacillus natto BN-NKPQQ-RWX-406 was obtained through genome rearrangement and high-throughput screening technology. The obtained NK and PQQ-producing Bacillus natto 10261 and 10263 were subjected to nitrosoguanidine mutagenesis respectively; then the offspring were mixed for mutagenesis, cytoplasmic fusion was carried out (fusogenic agent was polyethylene glycol 4000), subcultured, Realized genome shuffling, optimized and integrated the dominant mutations of all mutagenized progenies of Bacillus natto 10261 and 10623, and bred a high-yield NK and PQQ Natto with high yield, vigorous growth, stable genetics, and suitabl...

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Abstract

The invention discloses a method for breeding bacillus natto and lactobacillus casei shirota to ferment LactucapsativapL.var.asparaginapBailey. The method comprises the steps that two kinds of the bacillus natto 10261 and 10263 capable of producing nattokinase and pyrroloquinoline quinone are subjected to nitrosoguanidine mutagenesis, mutation progenies of the bacillus natto 10261 and 10263 are mixed for cell fusion subcultivation, a substrate induction adaptive strategy is further used on the basis of a genome rearrangement technique, and the bacillus natto containing rich nattokinase and pyrroloquinoline quinone is obtained after high-throughput screening; the lactobacillus casei shirota is subjected to nitrosoguanidine mutagenesis, and the lactobacillus casei shirota of methionine auxotroph is screened; the screened bacillus natto and lactobacillus casei shirota are fermented together and the LactucapsativapL.var.asparaginapBailey containing rich nattokinase and pyrroloquinoline quinone is obtained. The content of the pyrroloquinoline quinone in the fermented LactucapsativapL.var.asparaginapBailey containing rich nattokinase and pyrroloquinoline quinone reaches 88-107ng/mL, and the activity of the nattokinase reaches 875-1000U/mL.

Description

technical field [0001] The invention relates to the field of microbial engineering and fermentation, in particular to a method for co-fermenting lettuce with Bacillus natto and Lactobacillus casei. Background technique [0002] Natto is fermented from soybeans by Bacillus natto, and has multiple physiological functions: [0003] ①The effect of natto on the cardiovascular system: Nattokinase (Nattokinase, NK), a symbolic physiologically active substance in natto, is a protein capable of dissolving thrombus. It consists of 275 amino acid residues and has a molecular weight of about 27KD. It is non-toxic, It has no side effects and is a serine protease whose thrombolytic effect even exceeds that of the thrombolytic agent urokinase. In addition to its direct thrombolytic effect, nattokinase also has the ability to promote the production of plasminogen activator in venous endothelial cells, thereby indirectly expressing its thrombolytic activity. Linoleic acid in natto accounts...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A23L1/212C12N15/03C12N15/01C12R1/07C12R1/245A23L19/00
CPCC12N15/01C12N15/03A23V2002/00A23V2400/125A23V2200/30
Inventor 吴渊李青青张哲滔
Owner HANGZHOU YUANPEITE BIOTECH
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