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Method for detecting lactic acid bacteria in fermented dairy product

A technology for fermented dairy products and detection methods, applied in the field of detection of lactic acid bacteria in fermented dairy products, can solve the problems of time-consuming detection process, poor stability and reliability of detection results, etc., and achieve the effect of ensuring accuracy

Inactive Publication Date: 2015-01-14
中山鼎晟生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The traditional qualitative detection methods of lactic acid bacteria use physiological and biochemical reactions and selective medium for isolation and culture counting, but these methods are easily affected by various subjective and objective factors such as culture conditions and strain characteristics, the detection process takes a long time, and the stability of the detection results is low Reliability is poor, therefore, it is limited to use it for large-scale random sampling of commercial fermented dairy products

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] A method for detecting lactic acid bacteria in fermented milk products, comprising the following steps:

[0042] (1) Bacterial culture: Take 5ml of sample and add 10-20ml of sterilized water to dilute, shake and mix well, place a flat smear in MRS medium, incubate at 35°C for 12 hours, and take the sample colony;

[0043] (2) Take the colonies taken in step (1) for Gram staining, and observe the bacterial morphology under a microscope;

[0044] (3) Add the colony in step (1) to 10ml PBS buffer to make a bacterial suspension, take 1.5ml of the bacterial suspension and add it to a centrifuge tube, add an equal volume of chloroform, mix for 2min, centrifuge at 8000r Centrifuge at 8000r / min for 5min, take the supernatant, add sterile water or TE buffer 500ul to mix and resuspend, centrifuge at 8000r / min for 5min, take the supernatant as the sample solution to be tested;

[0045] (4) DNA extraction: Take the sample solution to be tested prepared in step (3) into another cen...

Embodiment 2

[0055] A method for detecting lactic acid bacteria in fermented milk products, comprising the following steps:

[0056] (1) Bacterial culture: Take 10ml of sample and add 20ml of sterilized water to dilute, shake and mix well, smear on the plate in MRS medium, incubate at 40°C for 24 hours, and take the sample colony;

[0057] (2) Take the colonies taken in step (1) for Gram staining, and observe the bacterial morphology under a microscope;

[0058] (3) Add the colony in step (1) to 20ml of PBS buffer to make a bacterial suspension, take 1.5ml of the bacterial suspension and add it to a centrifuge tube, add an equal volume of chloroform, mix for 5 minutes, and centrifuge at a speed 10000r / min, centrifugation time is 10min, take the supernatant, add sterile water or TE buffer 700ul to mix and resuspend, centrifuge, centrifugation speed is 10000r / min, centrifugation time is 10min, take the supernatant as the test sample solution;

[0059] (4) DNA extraction: Take the sample so...

Embodiment 3

[0068] A method for detecting lactic acid bacteria in fermented milk products, comprising the following steps:

[0069] (1) Bacterial culture: Take 8ml of the sample and add it to 15ml of sterilized water to dilute and oscillate to mix evenly, place a plate smear in MRS medium, incubate at 38°C for 18 hours, and take the sample colony;

[0070] (2) Take the colonies taken in step (1) for Gram staining, and observe the bacterial morphology under a microscope;

[0071] (3) Add the colony in step (1) to 15ml PBS buffer to make a bacterial suspension, take 1.5ml of the bacterial suspension and add it to a centrifuge tube, add an equal volume of chloroform, mix for 3 minutes, and centrifuge at a speed 9000r / min, centrifugation time is 8min, take the supernatant, add sterile water or TE buffer 600ul to mix and resuspend, centrifuge, centrifugation speed is 9000r / min, centrifugation time is 8min, take the supernatant as the test sample solution;

[0072] (4) DNA extraction: Take th...

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PUM

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Abstract

The invention relates to a method for detecting lactic acid bacteria in a fermented dairy product. The detection method comprises the following steps: firstly performing bacterial culture on a sample, further performing gram staining on a culture colony, observing the bacterial shape under a microscope, then preparing a bacterial suspension, centrifuging to prepare a sample solution to be detected, further performing DNA (deoxyribonucleic acid) extraction, finally performing PCR (polymerase chain reaction) amplification and electrophoresis detection, observing a detection result, judging that the sample is positive if a unique PCR amplification strip appears in the electrophoresis detection, judging that the sample is negative if other strips also appear when the strip appears, continuously performing illuminagenomeanalyzer sequencing on a PCR amplification product, and performing BLAST (basic local alignment search tool) comparative analysis on an obtained sequence by logging in an NCBI (National Center for Biotechnology Information) homepage. According to the detection method provided by the invention, the gram staining and the observation by the biological microscope are firstly performed, so that the lactic acid bacteria in the sample can be preliminarily screened, separated and identified; then PCR amplification and gel electrophoresis are further adopted for identifying the species of the lactic acid bacteria on the molecular level, and the detection result is accurate.

Description

[0001] technical field [0002] The invention belongs to the field of food detection, and in particular relates to a detection method for lactic acid bacteria in fermented milk products. Background technique [0003] All bacteria that can ferment carbohydrates to produce lactic acid are collectively called lactic acid bacteria. Lactic acid bacteria are a kind of Gram-positive bacteria. According to the different biochemical indicators formed during the evolution of their species, they can be divided into: low GC content group, such as Lactobacillus , Enterococcus and Lactococcus, etc., high GC-content bifidobacteria, these strains have different morphological structures, metabolic characteristics and physiological characteristics. Lactic acid bacteria naturally exist in human and other animals such as intestinal tract and vaginal mucosa. Lactic acid bacteria can help regulate the balance of intestinal microorganisms, promote nutrient absorption, improve liver function, prev...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/04C12R1/225C12R1/01
CPCC12Q1/686C12Q2531/113
Inventor 郭狄
Owner 中山鼎晟生物科技有限公司