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Synthetic medium for Staphylococcus carnosus, and preparation method and application of Staphylococcus carnosus fermentation broth

A technology for staphylococcus meatus and synthesizing culture medium, which is applied in the field of preparation of staphylococcus meatus synthetic culture medium and its fermentation liquid, can solve the problems of influence of separation and purification, many bands of miscellaneous proteins, differences, etc., and achieves reasonable formula and rich nutrition. , with a reasonable effect

Active Publication Date: 2017-12-22
TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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  • Application Information

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Problems solved by technology

[0005] For a long time, the compound medium used to cultivate Staphylococcus carnosus has complex components and rich carbon and nitrogen sources. Although the titer is high when using this compound medium for fermentation, due to the complex composition of the yeast extract, different origins , There are differences in different batches, which affect the quantitative and qualitative analysis of the experimental results, and when separating proteins, there are many bands of miscellaneous proteins, which have a great impact on the separation and purification of other substances

Method used

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  • Synthetic medium for Staphylococcus carnosus, and preparation method and application of Staphylococcus carnosus fermentation broth
  • Synthetic medium for Staphylococcus carnosus, and preparation method and application of Staphylococcus carnosus fermentation broth
  • Synthetic medium for Staphylococcus carnosus, and preparation method and application of Staphylococcus carnosus fermentation broth

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preparation example Construction

[0047] A method for preparing fermentation broth of Staphylococcus carnosus includes the following steps:

[0048] S1. Seed culture: The medium formula is (g / L): tryptone 10, yeast extract 5, sodium chloride 10; conditions: initial pH=6.5~8.0, culturing on a 180rpm shaker at 37℃ for 12h;

[0049] S2. Prepare the culture medium for growing Staphylococcus carnosus strain. The medium formula is (L): anhydrous glucose 10~50g, lysine 0.1~1.5g, valine 1.0~5.0g, glycine 0.1~1.5g, fine Amino acid 1.0~5.0g, proline 1.0~5.0g, glutamic acid 5~10.0g, tryptophan 0.1~0.5g, cystine 0.1~1.5g, thiamine hydrochloride 1.0~3.0mg, calcium pantothenate 1.0~3.0mg, niacin 1.0~3.0mg, manganese sulfate 1.0~3.0mg, potassium dihydrogen phosphate 0.1~1.0g, magnesium sulfate heptahydrate 0.1~0.8g, ferrous sulfate heptahydrate 0.005~0.010g, phosphoric acid trihydrate Dipotassium hydrogen 1.0~5.0g, 3-(N-morpholine)propanesulfonic acid 6~10g;

[0050] S3. Liquid fermentation culture: transfer the liquid seed solut...

Embodiment 1

[0052] Strain: Staphylococcus carnosus ( Staphylococcus carnosus ) ATCC 51365 / pBT2-ET - 5R - EGFP was constructed by the inventor's team, and its method is as follows:

[0053] Shuttle vector pBT2-ET-5R-EGFP: using pBT2 plasmid as the backbone, in its Kpn I and Nhe Insert one between the cloning sites tat - egfp The fusion gene is prepared; the promoter of the EGFP fusion gene is Staphylococcus carnosus ATCC 51365 eftu The promoter sequence of the gene (GenBank Accession No: 7551602), the signal peptide uses Staphylococcus carnosus ATCC51365 efeB The tat signal peptide sequence of the gene (GenBank Accession No: AM295250), and a linker containing 5 consecutive arginine sequences (5R) is added to the N-terminal of the subsequent EGFP gene (GenBank Accession No: AF302837); From the start codon of tat signal peptide to the base sequence reference of EGFP protein Figure 5 , Its base sequence is shown in SEQ ID NO. 1, and its amino acid sequence refers to Image 6 , As shown in S...

Embodiment 2

[0059] The composition of the fermentation synthesis medium used is (L): anhydrous glucose 40g, lysine 0.4g, valine 0.4g, glycine 0.4g, arginine 2.0g, proline 2.0g, glutamic acid 0.4g, Tryptophan 0.4g, cystine 0.4g, 1.0g, 1.5g, 2.0g, 3.0g, calcium pantothenate 2mg, niacin 2mg, thiamine hydrochloride 2mg, manganese sulfate 2mg, potassium dihydrogen phosphate 0.5g , 0.4g magnesium sulfate heptahydrate, 0.008g ferrous sulfate heptahydrate, 3g dipotassium hydrogen phosphate trihydrate, 8g 3-(N-morpholine) propanesulfonic acid.

[0060] First, the preserved Staphylococcus carnosus ( Staphylococcus carnosus ) ATCC 51365 / pBT2-ET - 5R - After the EGFP strain is activated to obtain a single bacteria, it is inoculated into a 250mL Erlenmeyer flask containing 50mL of seed culture medium, and continuously cultivated at a constant temperature of 180rpm in a 37℃ shaker for 12h, and then inoculates the bacterial solution to 50mL according to the 1% inoculum In the 250mL Erlenmeyer flask of the ...

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Abstract

The invention discloses a synthetic medium for Staphylococcus carnosus, and a preparation method and application of Staphylococcus carnosus fermentation broth. The synthetic medium for Staphylococcus carnosus comprises the following raw materials by weight: on the basis of a volume of 1 L, 10 to 50 g of anhydrous glucose, 0.1 to 1.5 g of lysine, 1.0 to 5.0 g of valine, 0.1 to 1.5 g of glycine, 1.0 to 5.0 g of arginine, 1.0 to 5.0 g of proline, 5 to 10.0 g of glutamic acid, 0.1 to 0.5 g of tryptophan, 0.1 to 1.5 g if cystine, 1.0 to 3.0 mg of thiamine hydrochloride, 1.0 to 3.0 mg of calcium pantothenate, 1.0 to 3.0 mg of nicotinic acid, 1.0 to 3.0 mg of manganese sulfate, 0.1 to 1.0 g of potassium dihydrogen phosphate, 0.1 to 0.8 g of magnesium sulfate heptahydrate, 0.005 to 0.010 g of ferrous sulfate heptahydrate, 1.0 to 5.0 g of dipotassium hydrogen phosphate trihydrate and 6 to 10 g of 3-(N-morpholino)propanesulfonic acid, with the balance being water. The invention also discloses a preparation method for the Staphylococcus aureus fermentation broth. The synthetic medium for Staphylococcus carnosus is reasonable in composition, rich in nutrients and capable of meeting the growth demands of Staphylococcus carnosus. The growth amount of Staphylococcus carnosus obtained through the synthetic medium provided by the invention is increased by 5% compared with the growth amount of Staphylococcus carnosus obtained through frequently used LB (Luria-Bertani) mediums.

Description

Technical field [0001] The invention belongs to the technical field of microbial culture medium and fermentation, and relates to a kind of Staphylococcus carnosus ( Staphylococcus carnosus ) Preparation method and application of synthetic medium and its fermentation broth. Background technique [0002] staphylococcus( Staphylococcus ) Is a gram-positive bacteria with an average diameter of 0.5-1.0μm. It is named after its stacked shape resembling a bunch of grapes and is generally not pathogenic. The representative species is Staphylococcus epidermidis ( Staphylococcus epidermidis ), Staphylococcus aureus ( Staphyloccocus aureus Rosenbach) and Staphylococcus saprophyticus ( Staphyloccocus saprophyticus )Wait. For a long time, people have used Staphylococcus for the fermentation of dried intestines. At first, they were considered to be micrococcus, but it turns out that these micrococcus were misclassified and are actually Staphylococcus. Based on DNA / DNA hybridization, bioc...

Claims

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Application Information

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IPC IPC(8): C12N15/74C12N15/62C12N15/66C12N1/20C12R1/44
CPCC07K14/43595C07K2319/02C12N1/20C12N15/74
Inventor 高强唐巧巧张变强朱燕
Owner TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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