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Method for quickly, qualitatively and quantitatively measuring Lactobacillus rhamnosus in probiotic dairy products

A technology for qualitative determination of Lactobacillus rhamnosus, applied in the direction of microbial-based methods, microbial determination/inspection, biochemical equipment and methods, etc., can solve the problems of time-consuming and labor-intensive, large impact, lack of stability of test results and problems Reliability and other issues

Inactive Publication Date: 2010-05-26
INNER MONGOLIA AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Traditional methods are easily affected by factors such as culture conditions and strain characteristics, and the detection efficiency is limited. Moreover, they are greatly affected by external environmental factors and medium performance. The detection results lack stability and reliability, and are time-consuming and labor-intensive.

Method used

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  • Method for quickly, qualitatively and quantitatively measuring Lactobacillus rhamnosus in probiotic dairy products
  • Method for quickly, qualitatively and quantitatively measuring Lactobacillus rhamnosus in probiotic dairy products
  • Method for quickly, qualitatively and quantitatively measuring Lactobacillus rhamnosus in probiotic dairy products

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0106] Rapid Qualitative Detection of Lactobacillus rhamnosus in Probiotic Dairy Products:

[0107] This embodiment uses the plasmid pRHT containing the target amplified fragment as a positive control; dH 2 O was used as a negative control; L. rhamnosus 1.2134, L. casei ATCC393, L. plantarum 1.2437, L. fermentum 1.188, L. acidophilus ATCC 4356, Bifidobacterium animalis Subsp. #, 36#, etc. are used as samples.

[0108] The L.rhamnosus 1.2134, L.plantarum 1.2437, and L.fermentum 1.188 were purchased from the China General Microorganism Culture Collection; L.acidophilus ATCC 4356 and L.caseiATCC393 were purchased from the American Type Culture Collection; L.casei Zhang and Bifidobacterium animalis Subsp.Lactis V9 is preserved by the Key Laboratory of "Dairy Biotechnology and Engineering" of the Ministry of Education of Inner Mongolia Agricultural University.

[0109] a. Preparation of template DNA

[0110] The sample DNA was extracted by the liquid nitrogen freeze-thaw-CTAB me...

Embodiment 2

[0125] Quantitative determination of Lactobacillus rhamnosus in probiotic dairy products:

[0126] Quantification of L. rhamnosus was performed on different samples mentioned in Example 1. Its determination steps are as follows:

[0127] a. Preparation of sample total RNA

[0128] Extraction by Trizol method: After grinding 500 mg of fermented milk products in a mortar filled with liquid nitrogen, quickly add 1 mL of Trizol reagent, extract sample RNA according to the instructions of the Trizol kit, and then treat it twice with DNaseI to ensure the complete elimination of DNA in RNA Contamination, the purified RNA sample was obtained, and the concentration was determined according to the method described in the specification of this application, and the RNA integrity was detected by 1% agarose gel electrophoresis.

[0129] b. Reverse transcription amplification

[0130] Reaction system 10.0 μL: 2.0 μL 5×PrimerScript Buffer (TakaRa DRR063A), 0.5 μL PrimerScript RT Enzyme Mix...

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Abstract

The invention relates to a method for quickly, qualitatively and quantitatively measuring Lactobacillus rhamnosus in probiotic dairy products. The method comprises the following steps: aiming at the Lactobacillus rhamnosus in probiotic dairy products, independently designing a species specificity primer of the 16S rRNA gene sequence of the Lactobacillus rhamnosus, carrying out species specificity PCR and real-time fluorescent quantitative PCR reaction by using the primer, and analyzing through an agarose gel electrophoresis pattern and a real-time fluorescent quantitative PCR pattern, thereby establishing the method for simply, conveniently, quickly, accurately, qualitatively and quantitatively measuring Lactobacillus rhamnosus in probiotic dairy products.

Description

【Technical field】 [0001] The invention relates to the technical field of dairy products containing probiotics. More specifically, the present invention relates to a rapid qualitative and quantitative determination method for Lactobacillus rhamnosus in probiotic milk products. 【Background technique】 [0002] Probiotic dairy products have become a hot topic due to their rich nutritional value and special beneficial effects. At present, there are many kinds of probiotics and dairy products containing probiotics on the market, and they are growing at a high rate every year. The supervision and certification of its quality, especially in terms of qualitative and quantitative detection, lacks scientific, reasonable and rapid methods. The qualitative and quantitative detection of probiotics in the traditional method still adopts the methods of physiological and biochemical reactions and selective medium pure culture isolation and counting. Traditional methods are easily affected...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N21/64C12R1/225
Inventor 张和平刘文俊包秋华
Owner INNER MONGOLIA AGRICULTURAL UNIVERSITY