菌種保藏的原則：
根據(jù)微生物生理、生化特點(diǎn)，人工創(chuàng)造條件，主要是低溫、干燥、缺氧三者，微生物的代謝處于不活潑、生長(zhǎng)繁殖受抑制的休眠狀態(tài)，以達(dá)到保持純種優(yōu)良性狀的目的。
常見菌種保藏方法介紹：
定期移植法（也稱傳代培養(yǎng)保藏法）
將菌種接種于適宜的培養(yǎng)基中，待生長(zhǎng)充分后，于4℃-6℃進(jìn)行保存并間隔一定時(shí)間進(jìn)行移植培養(yǎng)的菌種保藏方法。包括斜面培養(yǎng)、穿刺培養(yǎng)、液體培養(yǎng)等。
液體石蠟法（也稱礦物油保藏法）
是定期移植保藏法的輔助方法。將菌種接種到適宜的斜面培養(yǎng)基上，在最適條件下培養(yǎng)至菌種長(zhǎng)出健壯菌落后注入滅菌的液體石蠟，使其覆蓋整個(gè)斜面，再直立放置于4℃-6℃保存的一種保藏方法。
瓷珠保藏法
將培養(yǎng)好的微生物細(xì)胞或孢子制成懸浮液，轉(zhuǎn)入裝有無菌多孔玻璃珠（或瓷珠）的無菌瓶中，使其吸附于玻璃珠表面，去除多余的懸浮液，低溫冷凍保存的一種菌種保藏方法。
-80℃低溫保藏
將菌種保藏在-80℃冰箱的一種保藏方法。原理是低溫條件可減緩微生物的新陳代謝，以達(dá)到長(zhǎng)期有效的保藏微生物。
常見菌種保藏方法操作：
液體石蠟法（也稱礦物油保藏法）
將欲保存菌種接種到最適宜的培養(yǎng)基上；
選擇純凈優(yōu)質(zhì)石蠟油，經(jīng)121℃，30分鐘滅菌，滅菌后將其放烘箱中(160℃左右)干熱處理，除滅菌時(shí)滲入的水分；
待石蠟油變得清澈透明后，冷卻即可加入斜面，斜面上不能出現(xiàn)氣泡，液蠟添加量以高出斜面頂部約1cm左右為宜；
密封后保藏，保藏溫度要求在4～6℃。
瓷珠保藏法
將直接2mm-3mm的瓷珠分裝到小螺帽管中制成冷凍管，高壓滅菌；
標(biāo)記冷凍管，并在無菌條件下打開冷凍管；
將待保存菌株的新鮮培養(yǎng)物接入培養(yǎng)基中，制成菌懸液后，將菌懸液接入冷凍管中；
擰緊管帽，來回顛倒四到五次。勿使用渦旋振蕩器；
待菌株吸附到瓷珠上后，將多余菌懸液吸出。
-80℃低溫保藏
將培養(yǎng)好的菌種制成高濃度菌懸液(108～1010CFU/mL)；
取滅菌甘油與菌懸液充分混勻，使甘油濃度約為20%；
混勻后分裝入冷凍試管或安瓿中，于-80 ℃超低溫冰箱中保藏。
菌種保藏注意事項(xiàng)：
適宜的培養(yǎng)基（定期移植法與液體石蠟法）：
細(xì)菌--營養(yǎng)瓊脂；
真菌--PDA；
乳酸菌--MRS培養(yǎng)基；
厭氧菌--庖肉培養(yǎng)基；
嗜鹽性弧菌--含3%NaCl的TSA或營養(yǎng)瓊脂；
適宜的溫度：
根據(jù)菌株特性在適宜的溫度下保藏。如：副溶血弧菌、創(chuàng)傷弧菌、霍亂弧菌等致病弧菌，室溫(25℃以下)保存，不能冷藏。
適宜的氧氣條件：
根據(jù)菌株特性，提供適宜的氧氣條件，如微需氧條件或厭氧條件。
The principle of preservation of bacteria species:
According to the physiological and biochemical characteristics of microorganism, the conditions of artificial creation are mainly three cases of low temperature, dry and anoxic. The metabolism of microorganism is in a state of inactivity and inhibition of growth and reproduction, in order to keep the fine traits of pure species.
Introduction of common bacteria preservation methods:
Regular transplantation (also known as subculture preservation)
After inoculation, the strain was inoculated in suitable culture medium and stored at 4 -6 C for a long time. Including oblique culture, puncture culture, liquid culture and so on.
Liquid paraffin method (also known as mineral oil preservation)
It is an auxiliary method for the regular preservation of the preservation method. The bacteria were inoculated into the suitable slope culture medium, and under the optimum conditions, the liquid paraffin was incubated to the strain of the bacteria to infuse the sterile liquid, so that it covered the whole slope and was placed at 4 -6 C for preservation.
Preservation of porcelain beads
The cultured microbiological cells or spores are made into suspension and transferred into aseptic bottles containing sterile porous glass beads (or porcelain beads) to adsorb on the surface of glass beads and remove superfluous suspension and cryopreservation of a kind of bacteria preservation method.
Low temperature preservation at -80 C
A method of preserving bacteria in refrigerator at -80 C. The principle is that low temperature conditions can slow down the metabolism of microorganisms in order to achieve long-term and effective preservation of microorganisms.
Common bacteria preservation methods:
Liquid paraffin method (also known as mineral oil preservation)
Inoculate the desired species to the most suitable medium.
Choose pure and high quality paraffin oil, sterilize at 121 degrees, 30 minutes, and put it in dry oven (about 160 degrees Celsius) after sterilization.
When the wax oil becomes clear and transparent, it can be added into the inclined plane when cooling. The bubbles can not appear on the inclined surface. The amount of liquid wax added is about 1cm higher than the top of the slope.
After sealing, the storage temperature is required at 4~6 degrees C.
Preservation of porcelain beads
The direct 2mm-3mm ceramic beads were packed into a small nut tube and made into a freezing tube for autoclaving.
Mark the freezing pipe and open the freezing pipe under sterile conditions.
The fresh culture of the preserved strain was inserted into the culture medium, and the suspension was then made into the freezing pipe.
Tighten the cap and turn four to five times back and forth. Do not use a vortex oscillator;
After the bacteria were adsorbed on the porcelain beads, the superfluous bacteria suspension was sucked out.
Low temperature preservation at -80 C
The cultured strains were made into high concentration suspension (108 ~ 1010CFU/mL).
The glycerol concentration was about 20%.
After mixing, it is stored in a freezing test tube or ampoule and stored in a -80 low temperature refrigerator.
Precautions for the preservation of bacteria species:
Suitable medium (regular transplant method and liquid paraffin method):
Bacteria - nutritive agar;
Fungi --PDA;
The --MRS medium of lactic acid bacteria;
Anaerobes - Pao meat medium;
Vibrio halophilus (Vibrio halophilus) - TSA or nutritive agar containing 3%NaCl;
Suitable temperature:
It is preserved under suitable temperature according to the characteristics of the strain. For example, Vibrio parahaemolyticus, Vibrio vulnificus, Vibrio cholerae and other Vibrio at room temperature (below 25 degrees Celsius) can not be refrigerated.
Suitable oxygen conditions:
According to the characteristics of the strain, suitable oxygen conditions, such as micro aerobic or anaerobic conditions, are provided.
