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Bdellovibrio induced strain with strong cracking performance and wide cracking spectrum and application of bdellovibrio induced strain

A technique for mutagenic strains and Bdellovibrio, which is applied in the field of microbial mutation breeding, can solve the problems of difficult recovery and low mutation rate, and achieve the effects of enhancing lysis ability, improving lysis ability, and broadening the application range

Active Publication Date: 2020-07-03
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Traditional mutagenesis breeding is mainly based on ultraviolet mutagenesis, but its mutation rate is not high and requires multiple accumulated mutations, which is very easy to recover; and Co 60 Irradiation mutagenesis can obtain a higher mutation rate and a wider mutation spectrum, and it is not easy to recover
But through Co 60 There is no report on radiation mutagenesis to enhance the lytic performance of Bdellovibrio

Method used

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  • Bdellovibrio induced strain with strong cracking performance and wide cracking spectrum and application of bdellovibrio induced strain
  • Bdellovibrio induced strain with strong cracking performance and wide cracking spectrum and application of bdellovibrio induced strain
  • Bdellovibrio induced strain with strong cracking performance and wide cracking spectrum and application of bdellovibrio induced strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Example 1 Co 60 once irradiated

[0067] (1) Host culture: Take the newly cultivated host bacteria Bacillus subtilis GIM1.136, burn the mouth of the bottle under the outer flame of the alcohol lamp in the aseptic operation table, absorb 3mL of the host bacteria seed liquid and transfer it into a 250mL bottle of NB liquid In the culture medium, put the inoculated host bacteria in a constant temperature shaker at about 31°C for about 15 hours, then put them in a refrigerated centrifuge at 6000rpm and centrifuge at 4°C for 10min, then discard the supernatant, add to each tube 1mL of sterile distilled water with a salinity of 20‰, dispersed and mixed evenly, then transferred into a centrifuge tube to obtain the host bacteria concentrate, bagged, stored at 4°C, and used for later use;

[0068] (2) Preparation of 28 strains of host bacteria concentrate: Take one of each of the 28 strains of ampoule-preserved strains, wipe the mouth of the bottle with alcohol cotton, gently b...

Embodiment 2

[0082] Example 2 Co 60 secondary irradiation

[0083] (1) Host culture: same as step (1) of Example 1.

[0084] (2) Preparation of concentrated solution of 28 strains of host bacteria: same as step (2) of Example 1.

[0085] (3) BDE-1F1 seed liquid culture: Referring to step (3) of Example 1, pick phage plaques from the double-layer plate of the mutant strain BDE-1F1.

[0086] (4) Preparation of irradiated samples: take the BDE-1F1 seed solution in step (3), refer to step (4) of Example 1, and increase the irradiation dose to 5Gy / min×100min.

[0087] (5) Screening of mutagenic strains: Refer to step (5) of Example 1 to obtain the seed solution of the secondary mutagenic strain BDE-1F2.

[0088] (6) Detection of the ability of the mutant strain to lyse pathogenic Vibrio: refer to step (6) of Example 1.

[0089] Double-layer plate diagram of Bdellovibrio mutagen BDE-1F2 lysing Vibrio alginolyticus 2, see image 3 .

[0090] Two-layer plate diagram of Serratia figum 15 lyse...

Embodiment 3

[0103] Example 3 Co 60 Comparison of the ability of lysing positive bacteria between the mutant strain and the wild strain after the first and second irradiation

[0104] (1) Preparation of Gram-positive bacteria concentrate: Take one of the four strains of Gram-positive bacteria preserved in ampoules, wipe the mouth of the bottle with alcohol cotton, gently break the mouth of the bottle, and absorb 1 mL of NB liquid medium Pipette the powder until dissolved, transfer it to 50mL bottled NB liquid medium, incubate at 37°C, 200r / min for 8h, put it in a refrigerated centrifuge at 6000rpm, 4°C for 10min, then discard the supernatant, Add 1mL DNB liquid medium to each tube, mix well and transfer to a centrifuge tube to obtain a concentrated solution of Gram-positive bacteria, bag it, store at 4°C, and set aside;

[0105] (2) Detection of the ability of the mutagenic strain to lyse Gram-positive bacteria: refer to step (6) of Example 1.

[0106] Compare Co 60 The number and size ...

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Abstract

The invention discloses a bdellovibrio induced strain with strong cracking performance and a wide cracking spectrum and application of the bdellovibrio induced strain. The bdellovibrio induced strainis Bdellovibrio sp.BDE-1F2, is preserved on November 22, 2018 in Guangdong Microbiological Culture Collection Center and has a preservation number of GDMCC NO:60491. The cracking performance of bdellovibrio is enhanced by virtue of a Co60 irradiation induced mutation method, so that the cracking capability of the bdellovibrio to Gram negative and positive pathogenic bacteria and / or potential pathogenic bacteria, the application range of the bdellovibrio is broadened, and demands on production practice are better satisfied. Verification on the cracking performance of the bdellovibrio induced strain proves that four Gram negative bacteria which cannot be cracked in the past can be cracked, and the total cracking rate of the bdellovibrio to 28 indicator bacteria reaches up to 100%, the cracking capability of the bdellovibrio to the Gram negative bacteria is enhanced, and a method foundation is provided for enhancement of the cracking capability of the bdellovibrio.

Description

technical field [0001] The invention belongs to the technical field of microbial mutation breeding, in particular to Co 60 The application of radiation mutagenesis in enhancing the lytic performance of a strain of Bdellovibrio with Bacillus subtilis as a parasitic host, especially relates to a mutagenic strain of Bdellovibrio with strong lytic performance and wide lytic spectrum and its application. Background technique [0002] Bdellovibrio is a class of small bacteria capable of lysing other bacteria. It can parasitize and lyse Gram-negative bacteria of most families and genera. Its lysing effect on some pathogenic bacteria makes it of great value. At present, most Bdellovibrio cultures use Gram-negative bacteria as hosts, such as Escherichia coli. [0003] The growth and reproduction of Bdellovibrio is also affected by various factors, including the host. First of all, there must be a sufficient number of host bacteria to ensure the collision and adhesion of Bdellovibri...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A01N63/20A01P1/00C12R1/01
CPCC12N1/20C12N1/205C12R2001/01
Inventor 蔡俊鹏曹青青
Owner SOUTH CHINA UNIV OF TECH
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