Lactobacillus plantarum boosting immunological competence
A Lactobacillus plantarum, immune-enhancing technology, applied in the biological field, can solve problems such as large differences in probiotic activity
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Embodiment 1
[0028] Embodiment 1: Isolation and cultivation of bacterial strain
[0029] Collect traditional Chinese kimchi, grind it, dilute it with sterile normal saline gradient to an appropriate concentration, spread it on the MRS plate medium by the plate coating method, and culture it statically at 37°C for 36-72 hours to obtain Lactobacillus Individual colonies of suspected strains. The colonies were spread on the MRS plate medium, and anaerobically cultured at 37°C for 36-72 hours, and a single colony was picked for further purification. The strain identification was carried out according to the following Example 2, and the isolated strain BC299 was obtained.
[0030] Inoculate the isolated strains on the MRS plate medium by streaking method, culture anaerobically at 37°C for 36-72 hours, pick a single colony into 0.5-2mL fresh sterile MRS liquid medium, and incubate at 37±2°C Anaerobic culture for 24-48 hours, after the strain grows well, inoculate 5-8mL fresh sterile MRS liquid ...
Embodiment 2
[0031] Embodiment 2: bacterial strain identification
[0032] 2.1. Preliminary Analysis
[0033] Carry out preliminary analysis according to the standard method of strain identification, the result shows that isolated bacterial strain BC299 of the present invention is Gram-positive, catalase-negative bacillus (attached figure 1 ), no motility, no spore production, and can grow under both aerobic and anaerobic conditions.
[0034] 2.2 PCR analysis of 16S rDNA
[0035] Using the genomic DNA of strain BC299 as a template, add bacterial universal primers (P1 sequence: 5'-AGAGTTTGATCCTGGCTCAG-3'; P2 sequence: 5'-AAGGAGGTGATCCAGCC-3'), DNA polymerase, buffer, dNTPs to the PCR tube After waiting for the reagents, perform PCR amplification.
[0036] The amount of each reagent added and the amplification procedure are as follows:
[0037] Addition amount of each reagent: DNA genome (100ng / μL) 1μL as PCR reaction template, 0.5mmol / L primers P1 and P2 each 1μL, 10×PCR buffer (contain...
Embodiment 3
[0046] Embodiment 3: the acid resistance and bile salt resistance test of Lactobacillus plantarum BC299
[0047] In order to test whether Lactobacillus plantarum BC299 can successfully pass through the acidic and high bile salt environment of the gastrointestinal tract, so as to successfully exert its function of improving immunity in the intestinal tract, the experimental procedure is as follows:
[0048] After continuous activation of Lactobacillus plantarum BC299 in MRS liquid medium until rejuvenated, inoculate 4% (v / v) inoculum in MRS liquid medium with a pH value of 2.0, and perform static anaerobic culture at 37°C. Sampling was carried out at 0, 30, 60, 90, and 120 minutes respectively, and the gradient was diluted to 10 -5 ~10 -7 Afterwards, colony counting (mean ± SD) was carried out by the plate coating method to determine the tolerance of the strains to gastric acid.
[0049] In addition, the activated Lactobacillus plantarum BC299 was inoculated in the MRS liquid...
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