a vitamin b 12 strains and applications

A vitamin and B12 technology, applied in the field of bioengineering, can solve unseen problems and achieve the effect of stabilizing genetic traits

Active Publication Date: 2020-03-27
SHANGHAI INST OF QUALITY INSPECTION & TECHN RES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the existing literature, there is no information about Lysinibacillus producing vitamin B 12 report

Method used

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  • a vitamin b <sub>12</sub> strains and applications
  • a vitamin b <sub>12</sub> strains and applications
  • a vitamin b <sub>12</sub> strains and applications

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Primary screening of embodiment 1 isolate

[0027] (1) Obtaining of isolates

[0028] Perform a 10-fold gradient series coefficient on the fermented bean curd sample, take 100 μL of the dilution and pour it on the MRS plate, coat the plate and wait for the plate to dry. After culturing at 36°C for 3 days, colonies with different morphological characteristics are obtained on the plate, and different morphological colonies are picked from these plates. The isolated strains were transferred into MRS liquid medium and marked, and cultured at 36°C for 48h.

[0029] (2) Qualitative preliminary screening of isolates

[0030] Take part of the strain fermentation broth from each labeled test tube, mix it with distilled water at a ratio of 1:9 (that is, add 9 mL of distilled water to 1 ml of strain fermentation broth and mix well), then take 1 mL of the diluted solution, add 4 mL of distilled water and 5 mL of vitamin B12 For the test broth, sterilize at 121°C for 5 minutes. Af...

Embodiment 2

[0034] Example 2 isolates produce vitamin B 12 Verification and content determination

[0035] due to vitamin B 12 The qualitative screening and content determination are based on the microbiological method for the determination of vitamin B 12 On the basis of this method, for the stability and reliability of the results, the present invention adopts multiple methods to participate in bacterial strains to produce vitamin B 12 Verification of ability.

[0036] (1) Microbiological verification

[0037] pick vitamin B 12 In the qualitative results, isolates C5-1, C5-2, C5-8, C7-1 with high test tube turbidity and negative strains C8-2, C8-2 and L. plantarum ATCC 8014 were determined by microbiological method in the fermentation broth Vitamin B 12 content. Specific steps are as follows:

[0038] The isolates were inoculated into 10 mL of MRS broth and cultured at 36°C for 5 days. Take 5mL of fermentation broth and add 10mL of vitamin B 12 Pretreatment solution (1.3g of a...

Embodiment 3

[0050] Extraction and amplification of embodiment 3DNA

[0051] Using bacterial universal primers 27f and 1492r, the primer sequences are as shown in Table 2, the genomic DNA of the isolates C5-1, C5-8, and C7-1 verified in Example 2 were amplified by PCR, and the fragment size obtained from the amplification was 1.5kb fragments. Specific steps are as follows:

[0052] 1. Extraction of DNA

[0053] 1) Take 2-5mL bacterial liquid and centrifuge at 12000rpm for 5min, discard the supernatant;

[0054] 2) Add lysozyme (10mg / mL) to the precipitate, add 400μL, and bathe in 37℃ water for 1.5h;

[0055] 3) Add 400 μL CTAB and 20 μL proteinase K, and bathe in water at 56°C for 2 hours;

[0056] 4) After the water bath, add Tris-phenol:isoamyl alcohol:chloroform=25:24:1, 800μL, mix well, and centrifuge at 12000rpm for 10min;

[0057] 5) Take the supernatant (about 400 μL), add an equal volume of isoamyl alcohol: chloroform = 24:1, mix well, and centrifuge at 12000 rpm for 10 min; ...

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Abstract

The invention discloses a vitamin B-producing 12 strain, the preservation number of which is GDMCC No.60674. The invention also discloses the vitamin B-producing 12 The strain GDMCC No.60674 was used for the production of vitamin B 12 Applications. vitamin B 12 The strain GDMCC No.60674 can ferment and produce vitamin B through liquid static culture 12 , and the strain belongs to the genus Lysinibacillus, and there is currently no Lysinibacillus producing vitamin B 12 Therefore, the strain GDMCC No.60674 has good industrial application value.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, in particular, it relates to a vitamin B-producing 12 strains and their applications. Background technique [0002] Vitamin B 12 Also known as cobalamin (cobalamin), is a general term for a class of cobalt-containing corrin compounds, hydroxycobalamin (hydroxycobalamin), cyanocobalamin (cyanocobalamin), deoxyadenosylcobalamin (deoxyadenosylcobalamin) and Methylcobalamin is a vitamin B 12 main form of existence. Vitamin B 12 It is the most complex small molecular substance biosynthesized in nature. It participates in the metabolism of human cells, affects the synthesis and regulation of DNA, and also participates in the synthesis of fatty acids and the generation of energy. [0003] Since higher animals and plants cannot synthesize vitamin B 12 , although ESCHENMOSER et al completed vitamin B in 1973 12 The total chemical synthesis of (ESCHENMOSER A.Organische Naturstoffsynthese heut...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12P19/42C12R1/01
CPCC12P19/42C12N1/205C12R2001/01
Inventor 徐琼刘洋俞漪曲勤凤窦同海赵磊张娜娜钟江陈羽菲翁史昱
Owner SHANGHAI INST OF QUALITY INSPECTION & TECHN RES
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