Mycoplasma ovipneumoniae in-vitro culture medium and preparation method thereof

A technology for the preparation of mycoplasma pneumoniae and culture medium, which is applied in the field of bacterial culture medium and its preparation, can solve the problems of low culture medium bacteria quantity and poor efficiency, and achieve the effect of harmless to human body and simple production method

Inactive Publication Date: 2020-04-03
TARIM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Solve the problems of low bacteria culture and poor efficiency in the existing culture medium

Method used

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Examples

Experimental program
Comparison scheme
Effect test

example 1

[0013] 1. The preparation of culture medium of the present invention:

[0014] The chemical reagents used in the following examples are all conventional reagents and are commercially available.

[0015] Configure 1000mL ovine mycoplasma pneumoniae culture medium:

[0016] (1)PPLO Both 21g

[0017] (2) Glucose 10g

[0018] (3) Sodium pyruvate 10g

[0019] (4) 25% yeast extract 100mL

[0020] (5) 1% phenol red 3mL

[0021] (6) Sterile horse serum 200mL

[0022] (7) 50mg / mL ampicillin 1mL

[0023] Dissolve (1) to (5) reagents one by one into 700mL single distilled water and mix well, adjust pH to 7.8, autoclave at 115°C for 20 minutes, take out and cool to below 37°C. Add (6), (7).

[0024] 2. The configuration of the phenol red solution with a mass concentration of 1%:

[0025] Weigh 1.0g of phenol red, put it in a mortar, add 6mL (1mL each time) 1M NaOH (4.0g dissolved in 100mL single distilled water) in 6 times, grind until fully dissolved. Inhale the dissolved pheno...

example 2

[0029] 1. Comparative test

[0030] Preparation of KM2 medium:

[0031] MEM 5g, glucose 0.4g, sodium pyruvate 0.2g, hydrolyzed milk protein 5.1g, 1% phenol red 2.5mL, 25% fresh yeast extract 20mL, penicillin (200,000 IU / ml) 1mL, 10% thallium acetate 1mL, no Bacteria horse serum 200mL, mixed. Dilute to 1000ml with deionized water, adjust the pH value to 7.4 with sterilized 1M NaOH, filter and sterilize through a 0.22 micron filter membrane for later use.

[0032] 2. Detection

[0033] Viable bacterial titer (CCU) determination. The method is as follows: take 12 test tubes, add 4.5mL of the corresponding medium to each tube, add 0.5ml of a well-grown Mycoplasma ovis pneumoniae culture into the first tube, mix well with a shaker, replace the pipette with a new one, and start from the first tube. Pipette 0.5mL from tube 1 and add it to tube 2, then dilute 10 times to tube 11, and discard 0.5ml of culture solution in tube 11; the dilution of culture solution is 10 -1 -10 -11 ...

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Abstract

The invention discloses a mycoplasma in-vitro culture medium and a preparation method thereof. The preparation method of the mycoplasma in-vitro culture medium comprises the following steps: mixing PPLO (poly-p-phenylene oxide) Both, glucose, sodium pyruvate, a 25% yeast leaching solution and phenol red according to a certain ratio, and then carrying out autoclaving; after the temperature is stabilized at 37 or below, adding inactivated healthy horse serum and 50 mg / mL ampicillin dissolved by autoclaving water; and then adjusting the pH value to 7.6-7.8 by using 1 mol / L NaOH to obtain the culture medium. The mycoplasma in-vitro culture medium disclosed by the invention can be used for shortening the time required by culture and obviously improving the viable bacteria titer of mycoplasma ovipneumoniae, thereby laying a foundation for the development of high-quality vaccines of mycoplasma ovipneumoniae.

Description

technical field [0001] The invention relates to a bacterial culture medium and a preparation method thereof, in particular to a culture medium for Mycoplasma pneumoniae in vitro and a preparation method for the culture medium. Background technique [0002] Mycoplasma ovipneumoniae (Mycoplasma ovipneumoniae.MO) was first isolated from the lungs of sheep with lung adenocarcinoma by Mackay et al. (1963), a prokaryotic pathogenic microorganism without a cell wall between bacteria and viruses, which is the cause of sheep infection The pathogen of pleuropneumonia, sheep can manifest fever, cough, abdominal respiration, progressive emaciation and proliferative, suppurative pneumonia after respiratory infection, the incidence rate is 20% to 30%, and the fatality rate is generally 30% to 50%. , Some are as high as 80%, causing a large number of deaths of sheep, especially lambs, causing serious harm to the sheep industry. In my country, Mycoplasma ovis pneumoniae was first isolated ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12R1/35
CPCC12N1/20
Inventor 武军元杜奕州郗宇
Owner TARIM UNIV
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