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A technology of Bacillus coagulans and a screening method, applied in the field of microorganisms, can solve the problems of long cycle, low screening accuracy, heavy screening work, etc., and achieve the effect of high accuracy
Inactive Publication Date: 2017-12-01
JIANGSU WECARE BIOTECHNOLOGY CO LTD
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The screening methods in the prior art are mostly based on their biological and functional characteristics. These methods can be used for preliminary screening, but the screening work is heavy, the cycle is long, and the screening accuracy is low
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Embodiment 1
[0022] This embodiment provides a method for separating and purifying Bacillus coagulans, comprising the following steps:
[0023] S1. Take 5-10 g of intestinal content of weaned piglets, put it into sterile physiological saline containing small glass beads 10 times as much as the sample, and shake it for 5-10 minutes to make it evenly shaken.
[0024] S2. Place the sample suspension in a water bath at 55°C-65°C for 5-10 minutes, inoculate 2 mL of the suspension into a soluble starch medium, and incubate at 45°C-50°C, 150-200 r / min for 30-36 hours.
[0025] S3. Take the cultured bacterial solution and carry out 10-fold gradient dilution, take an appropriate dilution gradient and spread it on a bromocresol purple glucose solid plate, and incubate at 45-50° C. for 36-48 hours. After the colonies grow out, pick the colonies with yellow halos, separate and purify them, and store them as numbers 1-5.
[0026] In this embodiment, according to the characteristics of Bacillus coagula...
Embodiment 2
[0028] This example provides a method for identifying Bacillus coagulans, which can be identified by PCR amplification or 16s RNA sequencing alignment, or by combining the two methods to improve the accuracy of identification. This embodiment only provides an identification method combining the two.
[0029] 1. The PCR amplification method comprises the following steps:
[0030] As shown in Table 1, design specific primers P1 and P2 for Bacillus coagulans, use P1 and P2 as primers for PCR amplification, reaction conditions: denaturation at 95°C for 30 s, annealing at 48°C for 30 s, extension at 72°C for 2 min, cycle 30 times .
[0031] Table 1 Specific primers and general primers for 16S rRNA
[0032]
[0033] The strains obtained in the previous screening were amplified by PCR with specific primers for Bacillus coagulans, and subjected to agarosegel electrophoresis. The results were as follows: figure 1 As shown, only strain 3 has a specific band, and the band size is ...
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Abstract
The invention relates to the technical field of microorganisms and in particular relates to a screening method of bacillus coagulans. The screening method comprises the following steps: S1, dissolving a sample of bacillus coagulans in sterile water, and treating the sample in a water bath of 55-65 DEG C for a while to obtain a mixed cultureaqueous solution; S2, inoculating a certain mixed cultureaqueous solution to a soluble starch culture medium, and performing culture for a period of time to obtain mixed bacterial liquid; S3, diluting the mixed bacterial liquid and coating the diluted mixed bacterial liquid to a bromcresol purple glucose solid panel, performing culture for a period of time, picking up bacterial colonies with yellow halos and separating and purifying the bacterial colonies; and S4, identifying the screened and picked bacterial colonies by means of a method of PCR amplification and / or 16s RNA sequencing comparison to obtain bacillus coagulans. The screening method of the bacillus coagulans provided by the invention is simple, efficient and high in accuracy.
Description
technical field [0001] The invention relates to the technical field of microorganisms, in particular to a screening method for bacillus coagulans. Background technique [0002] Bacillus coagulans (Bacillus coagulans) is a type of lactic acid-producing bacteria that can form spores. In addition to maintaining the intestinal micro-ecological balance, stimulating immunity, improving the health of the body, and improving the digestive function of humans and animals, lactic acidbacteria also have strong stress resistance, gastric acid resistance, dryness resistance, high temperature and pressure resistance and high temperature resistance due to their ability to produce spores. It is widely distributed in soil, plantrhizosphere, animal manure and many human foods due to its unique biological characteristics such as easy storage. [0003] The research on Lactobacillus has become a hotspot in the research and development of probiotic strains. However, in the development of Bacil...
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