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

A technology for the qualitative determination of Lactobacillus acidophilus, 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 and reliability of test results sexual issues

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

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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 ...

Method used

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

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Experimental program
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Embodiment 1

[0105] Rapid Qualitative Detection of Lactobacillus Acidophilus in Probiotic Dairy Products:

[0106] This embodiment uses the plasmid pAT containing the target amplified fragment as a positive control; dH 2 O was used as a negative control; in addition, L.acidophilus ATCC 4356, traditional fermented lactic yogurt samples collected from Tibet 25#, naturally fermented lactic yogurt samples collected from Tibet 36#, L.casei ATCC393, L.rhamnosus 1.2134 and L.plantarum were used 1.2437 was performed as a sample.

[0107] The L. rhamnosus 1.2134 and L. plantarum 1.2437 were purchased from China General Microorganism Culture Collection; L. acidophilus ATCC 4356 and L. casei ATCC 393 were purchased from American Type Culture Collection.

[0108] a. Preparation of template DNA

[0109] The sample DNA was extracted by the liquid nitrogen freeze-thaw-CTAB method, and the specific steps were as follows:

[0110] (1) Take 0.5g probiotic fermented milk sample or lactic acid bacteria cul...

Embodiment 2

[0123] Quantitative determination of Lactobacillus acidophilus in probiotic dairy products:

[0124] Quantitative determination of NCFM on commercial Lactobacillus acidophilus. Its determination steps are as follows:

[0125] a. Preparation of sample total RNA

[0126] Trizol method extraction: 500 mg of fermented milk product was ground in a mortar filled with liquid nitrogen, and 1 mL of Trizol reagent was added quickly, and the sample RNA was extracted according to the instructions of the Trizol kit, and then treated twice with DNaseI to ensure that the DNA in the RNA was completely eliminated 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 integrity of the RNA was electrophoresed through 1% agarose gel.

[0127] b. Reverse transcription amplification

[0128] Reaction system 10.0 μL: 2.0 μL 5×PrimerScript Buffer (TakaRa DRR063A), 0.5 μL PrimerScr...

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Abstract

The invention relates to a method for quickly, qualitatively and quantitatively measuring Lactobacillus acidophilus in probiotic dairy products. The method comprises the following steps: aiming at the Lactobacillus acidophilus in probiotic dairy products, independently designing a species specificity primer of the 16S rRNA gene sequence of the Lactobacillus acidophilus, 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 acidophilus 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 acidophilus 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 affect...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N21/64C12R1/23
Inventor 张和平包秋华陈永福
Owner INNER MONGOLIA AGRICULTURAL UNIVERSITY
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