A kind of induction method of lactobacillus vbnc state

A technology of Lactobacillus and Lactobacillus casei, applied in the field of biology, can solve problems such as rare research

Active Publication Date: 2018-12-28
INNER MONGOLIA AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, there are few studies on the VBNC state of Lactobacillus derived from milk

Method used

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  • A kind of induction method of lactobacillus vbnc state
  • A kind of induction method of lactobacillus vbnc state
  • A kind of induction method of lactobacillus vbnc state

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0116] Embodiment 1: the induction of Lactobacillus casei Zhang VBNC state

[0117] The implementation steps of this embodiment are as follows:

[0118] A. Activation of Lactobacillus

[0119] The cryopreserved Lactobacillus casei Zhang was inoculated into the liquid MRS medium, activated and cultured at 37°C for 18 hours, and subcultured for three generations to obtain activated Lactobacillus casei Zhang, which was sold under the trade name Centrifuge 5810R by Eppendorf, Germany. Centrifuge, centrifuge at a speed of 10000r / min for 5min, according to the ratio of bacterial sludge in grams to sterile saline in milliliters is 1:3, the collected sludge is washed twice with sterile saline, the After each washing, the bacteria sludge needs to be centrifuged at a speed of 10000r / min for 3 minutes, and then the washed bacteria sludge is resuspended in the sterilized physiological In salt water, make Lactobacillus casei bacterial suspension;

[0120] B. Induction of Lactobacillus V...

Embodiment 2

[0137] Embodiment 2: the induction of Lactobacillus casei Zhang VBNC state

[0138] The implementation steps of this embodiment are as follows:

[0139] A. Activation of Lactobacillus

[0140] The cryopreserved Lactobacillus casei Zhang was inoculated into the liquid MRS medium, activated at 37°C for 24 hours, subcultured for three generations, and activated Lactobacillus casei Zhang was obtained, using a centrifuge sold under the trade name Centrifuge 5810R by Eppendorf, Germany. Machine, centrifuged at a speed of 10000r / min for 10min, according to the ratio of bacteria sludge in grams to sterilized saline in milliliters is 1:4, the collected bacteria sludge was washed 3 times with sterilized normal saline, and the bacteria After each washing, the sludge needs to be centrifuged at a speed of 10000r / min for 3 minutes, and then the washed bacteria sludge is resuspended in the sterile normal saline according to the ratio of bacteria sludge in grams to sterile saline in millilit...

Embodiment 3

[0155] Embodiment 3: the induction of Lactobacillus casei Zhang VBNC state

[0156] The implementation steps of this embodiment are as follows:

[0157] A. Pretreatment of Lactobacillus VBNC State Induction

[0158] The cryopreserved Lactobacillus casei Zhang was inoculated into the liquid MRS medium, and activated for 22 hours at a temperature of 37°C, subcultured for three generations to obtain the activated Lactobacillus casei Zhang, which was sold under the trade name Centrifuge 5810R by Eppendorf, Germany. Machine, centrifuged at a speed of 10000r / min for 8min, according to the ratio of bacteria sludge in grams to sterilized saline in milliliters is 1:5, the collected bacteria sludge was washed twice with sterilized normal saline, and the bacteria After each washing, the sludge needs to be centrifuged at 10000r / min for 3 minutes, and then the washed bacteria sludge is resuspended in the sterile normal saline according to the ratio of bacteria sludge in grams to sterile s...

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Abstract

The invention relates to a lactobacillus VBNC state induction and detection method. The method comprises activation of lactobacilli, induction of a lactobacillus VBNC state, detection of a lactobacillus VBNC state, fluorescence microscopy detection sample pretreatment and fluorescence microscopy detection. Lactobacillus casei Zhang is induced through sterilizing physiological saline at 4 DEG C so that the lactobacillus VBNC state is obtained. Lactobacillus bulgaricus ND02 is induced in a liquid MRS at 4 DEG C and then forms a VBNC state, and the VBNC state cell surface of the lactobacillus bulgaricus produces wrinkle so that length is reduced and the complete cell structure is kept. The lactobacillus in the VBNC state has a certain activity and can be recovered to a cultivable state under suitable conditions so that the method lays a foundation for researching lactobacillus activity in production. Through combination of a fluorescent dye (SYTO-9 and PI) and Leica DM4000B upright fluorescence microscope, a method which utilizes a glass slide to replace a filter film and realizes convenient and rapid detection of cell activity is built.

Description

【Technical field】 [0001] The invention belongs to the field of biotechnology. More specifically, the present invention relates to a method for inducing the VBNC state of Lactobacillus. 【Background technique】 [0002] At present, it is believed that bacteria usually have three states: "live cultivable state", "live non-culturable state" and "dead state", and active bacteria should refer to the first two states. As early as the early 1980s, Xu Huaishu et al. (Survival and viability of nonculturable Escherichia coli and Vibrio cholerae in the estuarine and marine environment.) first proposed the concept of viable nonculturable (VBNC) bacteria. This kind of living non-culturable bacteria will be in an abnormal physiological state under adverse external conditions, and cannot grow and reproduce on conventional medium, but the bacteria still maintain a low level of metabolic activity and certain gene expression in this state. When the harsh environment is lifted, it will return ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N1/00C12R1/225C12R1/245
CPCC12N1/005C12N1/20
Inventor 包秋华张和平王俊国王亚利
Owner INNER MONGOLIA AGRICULTURAL UNIVERSITY
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