Bifidobacterium longum 070103 with effects of remarkably reducing blood sugar and blood fat by targeting glucokinase and application of bifidobacterium longum 070103

A Bifidobacterium longum, glucose and lipid metabolism technology, applied in Bifidobacterium longum 070103 and its application fields, can solve the problem of lack of probiotics, achieve the effect of lowering blood sugar, fasting blood sugar and insulin content, and broad market prospects

Active Publication Date: 2022-07-22
GUANGDONG INST OF MICROBIOLOGY GUANGDONG DETECTION CENT OF MICROBIOLOGY +2
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  • Abstract
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  • Application Information

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Problems solved by technology

As a new drug target, there a...

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  • Bifidobacterium longum 070103 with effects of remarkably reducing blood sugar and blood fat by targeting glucokinase and application of bifidobacterium longum 070103
  • Bifidobacterium longum 070103 with effects of remarkably reducing blood sugar and blood fat by targeting glucokinase and application of bifidobacterium longum 070103
  • Bifidobacterium longum 070103 with effects of remarkably reducing blood sugar and blood fat by targeting glucokinase and application of bifidobacterium longum 070103

Examples

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Embodiment 1

[0027] Example 1 Culture, identification and preservation of lactic acid bacteria and preparation of lactic acid bacteria fermented milk

[0028] A total of 87 strains of lactic acid bacteria were isolated from the food and feces of healthy people. The human fecal strains were isolated from the previous stage of the research group. Inoculate 3% (v / v) lactic acid bacteria in MRS broth and 3% (v / v) in TPY broth supplemented with 1‰ of sterile 0.1% vitamin K1 and 5 mg / mL hemin ) of bifidobacteria were cultured at a constant temperature of 37 °C anaerobic to the logarithmic phase of the bacteria (about 16 h for lactic acid bacteria, about 40 h for bifidobacteria), passaged and activated twice, and set aside.

[0029] Bacterial DNA was extracted using a bacterial DNA extraction kit (Mabio, CHINA), followed by PCR amplification using 2 × PCR mix (Dongshengbio, CHINA). PCR amplification primers used 16S rRNA gene universal primer, upstream primer 27 F 5'-AGAGTTTGATCCTGGCCTCA-3' and...

Embodiment 2

[0034] Example 2: In vitro screening of probiotic fermented milk with activated glucokinase

[0035] The amino acid sequence of human hepatic glucokinase (GenBank: AAB97682.1) was entrusted to Jinweizhi Company for gene synthesis, cloned into pET-28A vector, sequenced and transformed into BL21 E. coli. The recombinant His-tagged GK protein was overexpressed by expansion at 37°C for 4 h, followed by induction with 0.5 mM isopropyl β-D-1-thiogalactoside (IPTG) for 48 h at 18°C. E. coli BL21 cells were centrifuged at 9,000 × g for 20 min at 4 °C and resuspended in lysis buffer (1 M NaCl, 50 mM phosphate buffered saline, 70 mM imidazole, 1 mg / mL lysozyme, pH = 7.6) and store at -80°C. Cells were thawed, sonicated on ice using an ultrasonic cell disruptor, and centrifuged at 15,000 × g for 30 min at 4 °C. The supernatant was filtered through a 0.8 μm filter and placed in 5 mL of HisTrap TM FF protein purification column, eluted with elution buffer (150 mM NaCl, 50 mM phosphate ...

Embodiment 3

[0042] Example 3 Safety evaluation of Bifidobacterium longum genome and phenotype

[0043] The whole genome of 515 Lactobacillus strains was sequenced by Illumina Nextseq 550 next-generation sequencing and Nanopore MinION third-generation sequencing platform. Bacterial genomic DNA extraction method is the same as before. Next-generation sequencing was performed using the AMT Rapid DNA-Seq Kit for Illumina (CISTRO, CHINA), and the High Output v2.5 kit (Illumina, USA) was used for sequencing. The library was constructed using the Rapid Barcoding Sequencing Kit (Nanopore, UK) for third-generation sequencing, and then sequenced using the R9.4.1 chip (Nanopore, UK). Trimmomatic (v0.39) and Filtlong (v0.2.0) software were used for quality control of the disembarkation data, respectively, and then Unicycler (v0.4.8) software was used for assembly. The assembled Lactobacillus genomes were evaluated by Quast (v5.0.2) software for genome quality control, and Abricate (v0.8.13) softwar...

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Abstract

The invention discloses bifidobacterium longum 070103 capable of remarkably reducing blood sugar and blood fat by targeting glucokinase and application of the bifidobacterium longum 070103. The preservation number of the Bifidobacterium longum 070103 is GDMCC (China General Microbiological Culture Collection Center) NO: 62526, and the preservation number of the Bifidobacterium longum 070103 is CGMCC NO: 62526. The bifidobacterium longum 070103 is separated from intestinal flora of healthy people, when the bifidobacterium longum 070103 is used for fermenting milk, the bifidobacterium longum 070103 has an activation effect on human liver glucokinase in vitro, in addition, the strain is derived from excrement of the healthy people and has no toxic or side effect on the human body, and meanwhile, the whole genome sequencing analysis finds that the strain is free of toxic genes, so that the bifidobacterium longum 070103 can be used for fermenting milk. Compared with traditional medicines for treating glucose and lipid metabolism disorder metabolic diseases such as obesity and type 2 diabetes mellitus, the pharmaceutical composition has certain advantages. The strain can be used for preparing probiotic fermented milk and has a wide market prospect.

Description

technical field [0001] The invention relates to the field of fermented dairy products, in particular to Bifidobacterium longum 070103, which has the significant hypoglycemic and hypolipidemic effects of targeting glucokinase and its application. Background technique [0002] Metabolic syndrome is a clinical condition characterized by a cluster of metabolic risk factors, characterized by central obesity, dyslipidemia, impaired glucose metabolism, elevated blood pressure, and low levels of high-density lipoprotein. There are three or more abnormal metabolic indicators at the same time and it is identified as having metabolic syndrome. However, with the change of lifestyle and the diversification of diet, abnormal glucose and lipid metabolism and other metabolic diseases caused by high-fat and high-sugar diet are becoming more and more common. [0003] The gut microbiota is closely related to the host's diet and metabolic parameters. Excessive intake of glycolipids is associat...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12Q1/689C12N15/11A23C9/12C12R1/01
CPCC12N1/20C12Q1/689A23C9/1234A23V2400/533
Inventor 吴清平蒋同代京莎谢新强李滢丁郁王涓陈谋通薛亮叶青华吴诗古其会张菊梅庞锐张友雄
Owner GUANGDONG INST OF MICROBIOLOGY GUANGDONG DETECTION CENT OF MICROBIOLOGY
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