I can only briefly introduce the three wide-ranging questions you asked.
1. culture medium is the material and environmental basis for microbial growth and reproduction. The culture medium contains nitrogen source, carbon source, water, growth factor and so on. It can be divided into solid culture medium, liquid culture medium, natural culture medium, synthetic culture medium and selective culture medium. Different media have different functions, some are used to activate microorganisms, some are used to select microorganisms, and some are used to preserve microorganisms. According to the specific purpose and the types of microorganisms, different media are selected. In addition, the proportion of nutrients used in the selected culture medium depends on the specific needs. As for preservation methods, there are short-term preservation and long-term frozen preservation, and there are many preservation methods. The temporary storage you mentioned is the 4℃ tilted tube storage method, in which microorganisms are labeled on the tilted tube culture medium and then stored at 4℃. You need culture medium. There are several different ways to preserve glycerol. Some sterile water+bacteria and glycerol are mixed in different proportions to form glycerol preservation solution with concentration of 65,438+00% ~ 40%, and some culture medium (liquid culture medium)+bacteria and glycerol are mixed in proportion to form glycerol preservation solution with concentration of 65,438+00% ~ 40%, and then frozen at -20 degrees Celsius. The shelf life is usually several years. Therefore, culture medium is also needed. This is only a small part of many media. Let's start with the first question.
2, a little dizzy, the second question is more complicated. The dilution coating plate method you mentioned is mainly used for the separation and purification of bacteria. When the concentration of bacterial suspension is too high, dense colonies will appear when inoculated into the culture medium, and the colonies will overlap with each other, losing the significance of separation and purification. If the concentration of bacterial suspension is too low, the bacterial suspension you apply to the plate may not contain the bacteria you need. It is also impossible to isolate the target bacteria.
What concentration is suitable for you to say one step at a time? You don't know, so you can only dilute it with 10 times, 1000 times and 10000 times, and then coat it on the corresponding plate with each concentration of bacterial suspension, so as to ensure that there will be a suitable concentration and the bacteria you want can be separated and purified. Of course, if you dilute it properly at one time, it is also possible. If you can isolate pure single colonies, no one dares to deny you, but the success rate is not high, or it is difficult. This method is also quite troublesome, but dilution is also a very simple operation. I wonder if you can understand this?
The third question is a little easier to describe. Principle: Pure water will form ice crystals at -20 degrees Celsius, destroying cell walls and membranes and killing bacteria. Glycerol does not form ice crystals, but it can maintain the integrity of cell walls and membranes. At the same time, because 100% glycerol can't mix bacteria evenly, and it is too sticky and does not flow, it should be diluted into 10%-40% glycerol, and our laboratory generally uses the final concentration of 20% glycerol for seed conservation. Note that the final concentration is 20%, not 20%. If the concentration is lower than 15% or 10%, there will also be ice crystals, which will destroy cell walls and cell membranes. On the other hand, bacteria are bacteria, not higher organisms. They are very resistant to adversity. -20 degrees Celsius will not kill them, but will only inhibit their activities, so they stop growing and enter a state similar to dormancy, but when conditions are right, they will continue to reproduce. Of course, the strains stored at glycerol -20 degrees Celsius should be activated before use. Note that this is the activation medium I mentioned earlier. It is a nutritious environment, and its purpose is to awaken dormant bacteria and restore their vitality. This answers your question about using culture medium on the other hand. It also has the function of activating bacteria. Don't say-20 degrees Celsius can't kill bacteria. Some bacteria can survive at 100 degrees Celsius. Of course, such bacteria are extreme microorganisms.
Having said so much, I wonder if you have read it carefully and understood it. Don't make me bother typing so many words, just take a look. ~ ~ ~ ~ and if you understand it, I hope you can share it with me. Recently, I asked some questions about gene cloning, and I used a lot, so I am earning points now. . . Thank you.