Expanded culture technology for microbial strains.

Microbial fermentation technology channel news: the expansion of the cultivation of bacteria is the first process of fermentation production, the process is also called seed preparationSeed preparation should not only increase the number of bacteria, but more importantly, after seed preparation, high-quality production seeds for fermentation productionTherefore, how to provide high fermentation yield, stable production performance, sufficient quantity and not contaminated with other hybrids is the key to seed preparation process, the task of expanding seed cultivationthe larger the industrial production, the more seeds are needed for each fermentationFor small microbes to complete such a huge fermentation transformation task in a short period of time in a matter of tens of hours, a large number of microbial cells must be availableThe purpose of the expansion of the species is to provide a considerable amount of metabolically active seeds for each fermentation tank feedBecause the length of fermentation time and the size of the inoculation, the amount of vaccination is large, fermentation time is shortThe addition of a large number of mature bacteria into the fermentation tank will help to shorten the fermentation time, improve the utilization rate of the fermentation tank, and also help to reduce the chance of contaminationTherefore, the task of seed expansion culture, not only to get pure and strong bacteria, but also to obtain a vibrant, sufficient number of inoculated bacteriaFor the fermentation process of different products, it is necessary to decide the number of seed expansion culture according to the growth and reproduction speed of the bacteria, and in the production of antibiotics, the cell growth and reproduction of the line bacteria is slow, and the three-level seed is often used to expand the cultureGenerally 50 t fermentation tank more than three-stage fermentation, some even use four-stage fermentation, such as streptomycin productionSome enzymes are also produced in three stages of fermentationThe fermentation of glutamate and other amino acids is a bacterial species, growth and reproduction speed is very fast, generally using secondary fermentation, seed preparation processbacteria, yeast seed preparation is a process of increasing the number of cellsThe bevel medium of bacteria uses a limited carbon source and a nitrogen-rich formula, and beef paste and protein is commonly used as an organic nitrogen source The mostly bacterial culture temperature is 37 degrees C, a few are 28 degrees C, the bacterial body culture time is generally 1 to 2 days, the bacteria producing spores need to be cultured for 5 to 10 days The seed preparation of mold and linebacteria generally consists of two processes, namely, the preparation of spores producing large quantities of spores on a solid medium and the seed preparation process of producing a large number of mycelium in a liquid medium 1, spore preparation spore preparation is the beginning of seed preparation, is an important link in fermentation production The quality and quantity of spores have a significant effect on the growth, reproduction and fermentation yield of mycelium in the future The spore preparation process of different species has different characteristics (1) the preparation of gliobacteria spores spores of gliobacteria generally use agar oblique spores culture, the medium contains some suitable for the production of spores of nutrients, such as bran, pea juice, protein and some inorganic salts Carbon and nitrogen sources should not be too rich (carbon source is about 1%, nitrogen source is not more than 0.5%), carbon source rich easy to cause physiological acidic nutritional environment, is not conducive to the formation of line bacteria spores, nitrogen source rich is conducive to mycelium reproduction and not conducive to the formation of spores In general, drying and restricting nutrients can directly or indirectly induce spore formation The culture temperature of the bevel of the line-dingbacteria is mostly 28 degrees C, a few of which are 37 degrees C, and the incubation time is 5 to 14 days which generation of oblique spores is used to be connected to liquid culture, depending on the characteristics of the bacteria The use of female oblique spores into the liquid medium is beneficial to prevent the variation of the bacteria, the use of sub-slope spores into the liquid culture medium can save the amount of bacteria species There are two ways for strains to enter seed jars A spore-in-the-tank method, in which oblique spores are made into spores and suspended directly into seed tanks This method can reduce the difference between batch and batch, has the advantages of convenient operation, simple process, easy to control the quality of spores, and so on, spore in-tank method has become a direction of fermentation production Another method is the shaker mycelium into the tank method, which is suitable for some slow-growing line-out bacteria, the advantage of this method can shorten the seed culture time in the seed tank (2) the preparation of mold spores mold spores culture, generally to rice, millet, corn, bran, wheat grain and other natural agricultural products as a medium This is due to the fact that the nutrients in these agricultural products are more suitable for mold spores, and that the surface area of these media is large enough to obtain a large amount of spores Mold culture is generally 25 to 28 degrees C, the incubation time is 4 to 14 days 2, seed preparation seed preparation is the solid medium cultured spores or bacteria into the liquid medium cultured, so that it reproduces into a large number of mycelium or bacteria process The medium and other process conditions used in seed preparation should be conducive to spore germination, mycelium reproduction or bacterial proliferation (1) Shaker Seed Preparation certain spores germination and mycelium reproduction slow bacteria, spores need to be cultured through the shaker into mycelium and then into the seed tank, this is the shaker seed The shaker is the equivalent of a miniature seed tank, and its medium formulation and culture conditions are similar to those of the seed can the bottle seed into the can, often using the mother bottle, sub-bottle two-level cultivation, sometimes the mother bottle seed can also be directly into the can Seed media requirements are relatively rich and complete, and easy to be decomposed and utilized by bacteria, nitrogen source rich is conducive to mycelium growth In principle, a variety of nutrients should not be too thick, the concentration of the sub-bottle medium is slightly higher than the mother bottle, closer to the seed tank's medium formula (2) seed preparation seed tank seed preparation process, depending on the bacteria, generally divided into first-level seeds, second seeds and third-level seeds preparation Spores (or shaker mycelium) are connected to smaller seed tanks and cultured to form a large number of mycelium, such seeds are called primary seeds, the first-level seeds into fermentation tank, called secondary fermentation If the first seed is connected to the larger seed tank, after being cultured to form more mycelium, the prepared seed is called the second-level seed, the second-level seed is transferred to the fermentation tank for fermentation, called the third-level fermentation By the same token, fermentation using three-level seeds is called four-stage fermentation the series of seed cans is mainly determined by the nature of the bacteria species and the growth rate of the bacteria and the rational application of fermentation equipment The purpose of seed preparation is to form a certain quantity and quality of bacteria When spores germinate and bacteria begin to multiply, the bacteria volume is small and can be done in small tanks The purpose of fermentation is to obtain a large number of fermentation products The product is formed after the mass formation of the bacteria and reaches a certain growth stage, which needs to be carried out in a large fermentation tank At the same time, the production of bacteria of several fermentation products, the different growth stages of nutrition and culture conditions of the requirements are different Therefore, the two different purposes, process requirements of the biological process in a large tank, not only affect the yield of fermentation products, but also cause the waste of power and equipment The reduction of seed tank series is beneficial to the simplification of the production process and the control of the fermentation process, which can reduce the fluctuation of fermentation caused by abnormal seed growth three, seed culture seed culture requires a certain amount of seeds, in the appropriate medium, control certain cultivation conditions and training methods, so as to ensure the normal growth of seeds Industrial microbial culture method is divided into two types of static culture and ventilation culture, static culture method is about to grow the medium in the fermentation container, after inoculation, not through the air for cultivation The production of gas culture method to oxygen bacteria and both oxycodone bacteria, they grow the environment must be supplied to the air, in order to maintain a certain level of dissolved oxygen, so that the rapid growth and fermentation of bacteria, also known as good gas culture 1, surface culture surface culture method is a kind of aerobic static culture method For the medium state in the container, it is divided into liquid surface culture and solid surface culture Compared with the medium volume in the container, the larger the surface area, the easier it is to promote the transfer of oxygen from the gas-liquid interface to the medium The growth rate of this method bacteria is related to the depth of the medium, the larger the surface area of the unit volume, the faster the growth rate 2, solid culture solid culture is divided into shallow solid culture and deep solid culture, collectively known as quertal culture It originated from the traditional musicmaking technology unique to brewing and production in China Its most important feature is the high enzyme vitality of solid crank 3, liquid deep culture liquid deep seed tank from the bottom of the tank ventilation, the air into the stirring blades dispersed into tiny bubbles to promote oxygen dissolution This culture method, which is stirred by the bottom of the tank ventilation, is called the deep culture method relative to the surface culture method of oxygen dissolved by natural diffusion by the gas-liquid interface It is characterized by easy to choose the best culture conditionaccordingly according to the nutritional requirements of the production of strains for metabolism and the conditions of ventilation, stirring, temperature and hydrogen ion concentration in the medium in different physiological periods (1) the three control points of the deep culture operation (1) sterilization fermentation industry requires pure culture, so the medium must be heated and sterilized before seed culture So the seed tank has a steam jacket to heat and sterilize the medium and the seed tank, or the medium is sterilized by a continuous heating sterilizer and continuously delivered to the seed tank (2) temperature control the culture medium sterilized, cooled to the culture temperature for seed culture, because as the growth and reproduction of microorganisms will produce heat, stirring will also produce heat, so to maintain a constant temperature, in the jacket or coil through the cooling water circulation (3) ventilation, stirring air before entering the seed tank to remove the bacteria through an air filter, to make asteria air, and then by the bottom of the tank into, and then by stirring the air to disperse into tiny bubbles In order to extend the bubble retention time, a vortex can be generated by installing the bezel in the tank In addition to increasing dissolved oxygen, the purpose of stirring can make the microorganisms in the culture liquid evenly dispersed in the seed tank, promote heat transfer, and pH are full, and the added acid and alkali uniform dispersion and other (2) several deep culture methods (1) control culture method according to the changes within the tank grasp the changes in the state of the short-term time and the possible determination of the impact of environmental factors on microbial metabolic activity, and to achieve the optimal conditions for the control of the culture To this end, the data is obtained by the sensor of the determination of the state variable, the comprehensive analysis is carried out by the electronic computer, and the resultist is used as a feedback adjustment signal to control the environment (culture condition) in a given baseline This is called computer control culture It is now widely used for the fermentation of large open-air cans of beer (2) Carrier training method carrier culture method is born out of the culture of the method, and at the same time absorbs the advantages of liquid culture, is a new development of the training method in recent years The characteristicis is to replace solid matrix such as bran with natural or synthetic porous materials as a carrier of microorganisms, and nutrients can be strictly controlled At the end of fermentation, the carrier can be reused as long as the bacteria and the medium are squeezed out for extraction (3) two-step in the two-step liquid deep culture of the enzyme preparation, each step of the bacteria is the same and the culture conditions are different, because the microbial growth and the most suitable conditions for the production of enzymes are very different For example, adding glucose to a medium can greatly increase the growth of mycelium or mycelium, but it seriously hinders the synthesis of many enzymes Although the ventilation of the culture can promote the growth of microorganisms, but in most cases it inhibits the synthesis of enzymes In order to obtain the largest high-energy enzyme, it is necessary to develop a regulatory method, which requires both high unit enzyme vitality of cells and a large number of cells, that is to say, to create different conditions for the bacteria in various physiological periods The deep cultivation of two-step liquid is one of the concrete measures to realize this kind of adjustment The two-step method of enzyme production is characterized by the distinction between the growth condition (long-term) of the bacteria and the enzyme-producing condition (the production period) The bacteria first multiply on the rich medium, then collect the concentration of the bacteria, wash and then transfer to the enzyme-producing medium that adds the induction, during which the bacteria accumulate a large number of enzymes, generally no longer reproduce, nutrients or inducing substances are fully utilized , the control of seed quality seed quality is an important factor affecting the level of fermentation production The quality of seeds mainly depends on the genetic characteristics of the species itself and the conditions of culture This means that there must be excellent strains, but also have good training conditions in order to obtain high-quality seeds (i) the factors that affect the quality of spores and their control spores quality is related to the medium, culture temperature, humidity, culture time, inoculation, etc.,