CN103074244B - Method for breeding dominant strains by using streptomyces aureofaciens growth period conditions - Google Patents

Abstract

The present invention relates to a method for breeding dominant strains by using streptomyces aureofaciens growth period conditions. The method comprises: taking streptomyces aureofaciens at a vigorous growth period, carrying out plate separation culture, selecting predominant strains to carry out slant culture, and carrying out primary shake flask screening and rescreening identification to select the dominant strains, wherein the dominant strains are subjected to low temperature pure seed storage after biology stability of the dominant strains is determined through a plurality of rescreening, and the dominant strains are subjected to trial use in industrial production so as to achieve stabilize or increase industrial production capacity of the strains. According to the present invention, a series of the screening is performed to obtain the three strains such as RC2-88, RC2-185 and RC2-196, wherein titers and yields are improved to different degrees after the three strains are applied in production, and compared with the starting strain, the industrial production efficacy is increased by 1-16%; and the strain breeding method is enriched, the yield is increased, and significant economic benefits and social benefits are provided.

Description

A kind of method of utilizing the winning bacterial strain of streptomyces aureus condition in vegetative period seed selection

Technical field

The invention belongs to biological technical field, especially a kind of method of utilizing the winning bacterial strain of streptomyces aureus condition in vegetative period seed selection.

Background technology

Duomycin (Chlortetracycline), claims again Uromycin, belongs to the one of derivative in tetracyclines.Chemical name: 6-methyl-4-dimethylamino-3,6,10,12,12a-penta hydroxy group-1, the chloro-Isosorbide-5-Nitrae of 11-dioxo-7-, 4a, 5,5a, 6,11,12a-octahydro-2-tetracene methane amide.Molecular weight: 478.8; Molecular formula: C ₂₂h ₂₃clN ₂o ₈.Duomycin has a broad antifungal spectrum, all can produce restraining effect to positive bacteria, negative bacterium, spirochete, Rickettsiae, mycoplasma, chlamydozoan, some protozoon etc.The anti-microbial effect of duomycin is mainly the growth of anti-bacteria, has germicidal action in the time of its higher concentration, and its mechanism of action is the synthetic of interferencing protein.Be applicable to the diseases such as white dysentery, typhoid fever, pig bacterial enteritis and bird paratyphoid, cholera.Meanwhile, as desirable fodder additives, it also has prevention latent infection, improves digestion and metabolism, suppresses harmful microorganism in enteron aisle, promotes beneficial microorganism growth, improves feed usefulness etc.In the world, duomycin is the kind of usage quantity maximum in all feed medicated premixs, is one of important in the world microbiotic for animals.

Streptomyces aureus (Streptomyces aureofaciens) is Streptomycetaceae, streptomyces, and spore filament length, 5-8 encloses volution.Spore is spherical in shape, oval, Long Circle, and surface is slightly rugged and rough.It is golden yellow that bacterium colony is.Below 30 DEG C time, the ability of synthetic duomycin is stronger, in the time that temperature exceedes 35 DEG C, and only synthetic tsiklomitsin and synthetic duomycin.

The object of culture presevation is to keep as far as possible the stable of its original proterties and vigor, guarantees that bacterial classification is not dead, does not make a variation, not contaminated, so that research, exchange and use.In actual production practice process, bacterial classification is through long-term artificial culture or preservation, because appearring in the effect of spontaneous mutation, negative sudden change causes that some good characteristic dies down or disappears, the result of this variation often causes the degeneration of bacterial classification, and natural variation in streptomyces aureus production process tends to occur gain mutant, produce strain excellent.

Summary of the invention

The object of the invention is to utilize the winning bacterial strain of streptomyces aureus condition in vegetative period seed selection, provide a kind of spontaneous mutation taking microorganism as basic strain breeding method.

The object of the invention is to be achieved through the following technical solutions:

(1) producing prior fermentation liquid cultivates;

(2) plate separation and Culture;

(3) choosing single colony inoculation test tube slant cultivates;

(4) shaking flask primary dcreening operation, multiple sieve, the qualification of tiring;

(5) sand pipe is preserved;

(6) production application;

Advantage of the present invention and beneficial effect are:

1, the present invention utilizes the streptomyces aureus of producing early stage, is just sieved and obtains winning bacterial strain again by plate separation and Culture, test tube slant cultivation, shaking flask, and application is produced, and raising is tired and output, has significant economic benefit.Meanwhile, for strain improvement provides the new mode of thinking and method.

2, the present invention has determined method and the condition of utilizing streptomyces aureus to produce the winning bacterial strain of conditional filtering in early stage, for further researching and developing the resources of production, and is meeting livestock industry demand, is promoting the aspects such as feed industrial development that important realistic meaning is provided.

Embodiment

Below in conjunction with embodiment, the present invention is further described, and following embodiment is illustrative, is not determinate, can not limit protection scope of the present invention with following embodiment.

Embodiment 1:

A method of utilizing the winning bacterial strain of streptomyces aureus condition in vegetative period seed selection, the steps include:

(1) choose the streptomyces aureus fermented liquid of producing to 29h, be seeded to separation plate.

(2) plate separation and Culture: cultivate 3-5 days under 32 DEG C of conditions to ripe for subsequent use.

(3) test tube slant is cultivated: picking list bacterium colony from cultured plate, inoculation test tube slant,, cultivates 3-5 days by 32 DEG C.

(4) shaking flask primary dcreening operation, multiple sieve, the qualification of tiring: test tube slant is seeded to shake-flask seed substratum, cultivates 17h at 28 DEG C, inoculation Medium of shaking flask fermentation,, cultivates 7 days by 28 DEG C, survey is tired, select this batch to tire/normal produce tire >=105% or approach 105% bacterial strain and carry out multiple sieve.

(5) sand pipe is preserved.

(6) production application: RC2-196 is applied in production, finally tires and improved 5% and 16% than normal production batch respectively with total hundred million.

Embodiment 2:

A method of utilizing the winning bacterial strain of streptomyces aureus condition in vegetative period seed selection, the steps include:

(1) choose the streptomyces aureus fermented liquid of producing to 29h, be seeded to separation plate.

(2) plate separation and Culture: cultivate 3-5 days under 32 DEG C of conditions to ripe for subsequent use.

(3) test tube slant is cultivated: picking list bacterium colony from cultured plate, inoculation test tube slant,, cultivates 3-5 days by 32 DEG C.

(4) shaking flask primary dcreening operation, multiple sieve, the qualification of tiring: test tube slant is seeded to shake-flask seed substratum, cultivates 17h at 28 DEG C, inoculation Medium of shaking flask fermentation,, cultivates 7 days by 28 DEG C, survey is tired, select this batch to tire/normal produce tire >=105% or approach 105% bacterial strain and carry out multiple sieve.

(5) sand pipe is preserved.

(6) production application: RC2-185 is applied in production, finally tires and improved 6% and 1% than normal production batch respectively with total hundred million.

Claims (1)

1. a method of utilizing the streptomyces aureus fermented liquid of producing early stage to screen winning bacterial strain, is characterized in that, the steps include:

(1) get in production and ferment to the streptomyces aureus fermented liquid of 29h, be seeded to separation plate;

(2) plate separation and Culture: cultivate 3-5 days under 32 DEG C of conditions to ripe for subsequent use;

(3) test tube slant is cultivated: picking list bacterium colony from cultured plate, and inoculation test tube slant,, cultivates 3-5 days by 32 DEG C;

(4) shaking flask primary dcreening operation, multiple sieve, the qualification of tiring: test tube slant is seeded to shake-flask seed substratum, cultivates 17h at 28 DEG C, inoculation Medium of shaking flask fermentation,, cultivates 7 days by 28 DEG C, survey is tired, select this batch to tire/normal produce tire >=105% or approach 105% bacterial strain and carry out multiple sieve;

(5) the RC2-196 bacterial strain sand pipe of multiple sieve is preserved;

(6) production application: RC2-196 is applied in production, finally tires and improved 5% and 16% than normal production batch respectively with total hundred million.