Inoculation, isolation and culture of microorganisms

Summary

Microorganisms exist in a mixed state in nature, from which they must be isolated to obtain the desired strain. In the preservation of strains of bacteria accidentally contaminated also need to be purified. Microbial isolation and purification of many methods, but the basic principle is similar, that is, to be separated from the sample for a certain amount of dilution, and make the microbial cells (or spores) as much as possible in a dispersed state, and then make it grow into a single colony of pure species. However, the above work can not be separated from inoculation, i.e., the process of transferring one microorganism to another sterilized medium.

Operation method

Inoculation, isolation and culture of microorganisms

Principle

The inoculation and isolation of microorganisms and culture is one of the basic steps to do cell metabolism analysis and in vitro experiments, the inoculation and isolation of good and bad directly affect the culture of microorganisms as well as the subsequent experimental operations and results (1) for cell isolation (2) cell purification (3) cell proliferation culture.

Materials and Instruments

Escherichia coli, Bacillus subtilis, Staphylococcus aureus, yeast.
Lysol, Carbonic acid, Formalin, Beef paste, Peptone.
Thermostat incubator Inoculation ring Alcohol lamp

Move

1, microbial inoculation: refers to the pure species of microorganisms or bacteria-containing materials transferred to another nutrient substrate on the process. According to different purposes can be used for different inoculation methods, such as plate scribing method, coating method, pouring method.

2, microbial isolation: refers to the characteristics of microorganisms, using a variety of methods from the bacterial samples from a single individual (such as a single organism or spore) or a section of mycelium growth and reproduction of microbial groups formed by the process. Commonly used isolation methods include plate delineation method, dilution coating method, plate separation method, dilution pouring plate separation method and single cell separation method. The basic principle of plate separation method includes two aspects: (1) Selection of microorganisms to be separated from the growth conditions, such as nutrition, acidity and alkalinity, temperature and oxygen requirements or the addition of a certain inhibitor to cause the growth of the microorganisms only conducive to the environment. (2) Individual colonies formed by microorganisms growing on solid media can be aggregates propagated from a single cell, so that a pure culture can be obtained by picking single colonies. It is worth pointing out that from the microbial population growth on the formation of a single colony can not necessarily guarantee a pure culture.

3, microbial culture: refers to microorganisms are inoculated into the nutrient substrate, the process of growth and reproduction under certain conditions, divided into aerobic culture method and anaerobic culture method.

Caveat

1. During inoculation, it is necessary to operate next to an alcohol lamp in order to maintain a relatively sterile environment. The inoculation ring and test tube spout should not be left arbitrarily on the table or in contact with other objects.

2. The plate or slant needs to be scratched quickly and gently, without scratching the medium.

3. After inoculation, pull out the inoculation ring, cauterize the mouth of the tube, and put on the silicone plug. Do not meet the silicone plug with the mouth of the test tube when putting on the silicone plug, so as not to incorporate unclean air into the test tube in the flowing air.

4. When doing plate partitioning and delineation separation, after delineating a plot, uniformly and thoroughly cauterize the inoculation ring and sterilize it, and then delineate the next plot after it cools down.

5. All culture vessels need to be strictly sterilized.

Common Problems

1. A colony is a group of daughter cells visible to the naked eye formed by the growth and multiplication of a single or a few cells on the surface of a solid culture medium or inside it. The morphology of the colony may vary to different degrees depending on the strain or the same strain but with different conditions such as medium, incubation temperature, pH value and time. If the medium used, culture temperature and time conditions are the same, the colony morphology formed by the same bacteria has relative stability and uniformity.

2

. The determination of pure culture in addition to the observation of its colony characteristics, but also combined with the microscope to detect the individual morphology characteristics before deciding, some microorganisms pure culture after a series of isolation and purification process and the identification of diversity characteristics can be determined.

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