What are the Functions of Incubator in Biotechnology Field

What is Incubator?

Biological incubator is a kind of biological culture equipment with refrigeration and heating two-way temperature control system, and has temperature control function, is one of the indispensable laboratory equipment of plant, biology, microorganisms, genetics, viruses, environmental protection and other scientific research and teaching departments, which is widely used in constant temperature tests, culture experiments, environmental experiments and other fields, and plays a pivotal role in scientific research, environmental protection, sewage treatment and other departments.

With the rapid development of biomedicine, bacterial incubators are widely used in the field of biology, but environmental factors have a great impact on the culture of bacteria, each bacterium has different requirements for the culture environment, and some bacteria have high requirements or even harsh, slight changes can easily lead to culture failure, to experiments, research and other work to bring a lot of obstacles. At present, in real life, there is no convenient and easy-to-use bacterial culture device, and when bacterial culture is required, traditional bacterial culture devices are usually used, which is difficult to effectively and conveniently regulate the bacterial culture environment and affect the bacterial culture effect. In addition, it affects the progress of enterprise work and increases the difficulty of work for staff. But LIB INDUSTRY managed to solve this problem.

The Structure of LIB Incubators for Biological Lab

LIB incubators include:

(1)Culture shelf, the culture shelf is used to place the fungus bag;

(2)Temperature control internal fan, temperature control internal fan placed at the top of the culture frame;

(3)Exhaust device, used to provide oxygen during the growth of mycelium, exhaust device includes intake fan and exhaust fan, and the intake fan is the same as the exhaust fan and the temperature control inner fan;

(4)Constant temperature device, used to control the germination temperature in the environment; and purification devices for filtering and disinfecting the air in the germinated environment.

The structure of the bacterial culture system of LIB incubator is reasonably set to ensure the oxygen and temperature required for bacterial development, and improve the efficiency of bacterial culture; At the same time, the germination environment was filtered and disinfected to ensure the survival rate and quality of edible fungus culture.

Bacterial Culture

1.Staphylococcus aureus

Staphylococcus aureus is a Gram-positive spherically shaped bacterium, a member of the Bacillota, and is a usual member of the microbiota of the body, frequently found in the upper respiratory tract and on the skin. It is often positive for catalase and nitrate reduction and is a facultative anaerobe that can grow without the need for oxygen.

In humans, S. aureus can be present in the upper respiratory tract, gut mucosa, and skin as a member of the normal microbiota.However, because S. aureus can cause disease under certain host and environmental conditions, it is characterized as a "pathobiont".

High Temperature, High Salt

Staphylococcus aureus has a certain tolerance to high temperature, in a high temperature environment above 80 °C for 30min to completely kill it, in addition, Staphylococcus aureus can survive in a high-salt environment, up to 15% concentration of NaCl solution. Due to the structural characteristics of the bacteria themselves, 70% ethanol can quickly killed them within a few minutes. The metabolic type of Staphylococcus aureus is aerobic or facultative anaerobic, the environmental requirements are not high, 37 °C is the optimal growth temperature, can survive in a variety of harsh environments, therefore, the bacteria can be cultivated normally with general nutritional agar.

(1) The raw pig heart is added to 1 to 2 times of water, pulverized, filtered, and the first filtrate and filter residue are obtained;

(2) Add 1 to 2 times of water to the filter residue, soak at 90 to 95° C., and filter to obtain the second filtrate and filter residue; combine the first filtrate and the second filtrate to obtain the third filtrate;

(3) will calculate according to liquid gross weight, add 0.025～0.5g% peptone in the 3rd filtrate, and 0.3～0.9g% sodium chloride adds the 3rd filtrate, and in this filtrate, add 0.5% gac, will Its pH value is adjusted to about 7.2 to obtain a culture medium;

(4) resuscitating the Staphylococcus aureus bacterial strain, making seed liquid after enriching the bacteria, inoculating after making its bacterial concentration reach 105-107, its inoculum size is 0.02～0.2ml%;

(5) After about 15-20 hours of fermentation at 35°C, a culture was obtained,

(6) Sterilize the culture, filter it, adjust its osmotic pressure to isotonicity, and adjust its pH value to 6.8-7.4 to obtain the final product.

2.Escherichia Coli

Escherichia coli is a Gram-negative, facultative anaerobic, rod-shaped, coliform bacterium of the genus Escherichia that is commonly found in the lower intestine of warm-blooded organisms. Most E. coli strains are harmless, but some serotypes such as EPEC, and ETEC are pathogenic and can cause serious food poisoning in their hosts, and are occasionally responsible for food contamination incidents that prompt product recalls.

Optimum growth of E. coli occurs at 37 °C (99 °F), but some laboratory strains can multiply at temperatures up to 49 °C (120 °F). E. coli grows in a variety of defined laboratory media, such as lysogeny broth, or any medium that contains glucose, ammonium phosphate monobasic, sodium chloride, magnesium sulfate, potassium phosphate dibasic, and water. Growth can be driven by aerobic or anaerobic respiration, using a large variety of redox pairs, including the oxidation of pyruvic acid, formic acid, hydrogen, and amino acids, and the reduction of substrates such as oxygen, nitrate, fumarate, dimethyl sulfoxide, and trimethylamine N-oxide. E. coli is classified as a facultative anaerobe. It uses oxygen when it is present and available. It can, however, continue to grow in the absence of oxygen using fermentation or anaerobic respiration. Respiration type is managed in part by the arc system. The ability to continue growing in the absence of oxygen is an advantage to bacteria because their survival is increased in environments where water predominates.

Culture of E. coli:

37 °C, constant temperature culture for 48 to 72 hours, because of the difference between inoculation and specific culture conditions, its growth is generally 2 days away to grow obvious colonies.Colony characteristics: milky white, round, neat colony edges, smooth surface, consistent surface and back color.

3.Staphylococcus epidermidis

Staphylococcus epidermidis is a Gram-positive bacterium, and one of over 40 species belonging to the genus Staphylococcus. It is part of the normal human microbiota, typically the skin microbiota, and less commonly the mucosal microbiota and also found in marine sponges. It is a facultative anaerobic bacteria.

Staphylococcus epidermidis has low nutritional requirements and grows well on ordinary medium. Aerobic or facultative anaerobic, the optimum growth temperature is 37°C, and the optimum pH is 7.4. It is salt tolerant and requires biotin for growth. After 24-hour incubation at 37°C in broth medium, it grows uniformly turbid. Form raised, smooth opaque round colonies, 1-2 mm in diameter, on plain agar plates. Staphylococcus epidermidis generally produces white or lemon pigments, so colonies appear white or lemon.

Fungus Culture

1.Candida albicans

Candida albicans is an opportunistic pathogenic yeast[5] that is a common member of the human gut flora. It can also survive outside the human body.[6][7] It is detected in the gastrointestinal tract and mouth in 40–60% of healthy adults.

(1)Weigh 47g of dry powder, add 1000mL of distilled water, pressurize at 121°C for 15min, cool to 50°C, pour onto a plate, and set aside.

(2)Add 1000mL distilled water to 1000mL dry powder (10.0g peptone, 20.0g glucose, 2.0g mixed color reagent, 0.5g chloramphenicol, 15.0g agar), boil for 2min, cool to 50°C, pour plate, spare.

(3)Candida chromogenic medium: Take 3.0g of yeast powder, 3.0g of malt extract powder, 5.0g of peptone, 20.0g of glucose, 15g of agar, 0.05g of chloramphenicol, 2.0g of mixed chromogenic substrate, 1.0g of glycine, and heat to melt Afterwards, adjust the pH to 6.4, boil for 2 minutes, cool to 50°C, pour into a plate, and set aside.

(4)Inoculate Candida albicans ATCC10231 in yeast broth, culture at 35°C for 24-48h, then take a small amount of culture solution to make 102 bacteria suspension, then carry out 10-fold gradient dilution, take 0.1mL of bacteria solution and apply it to the display screen of the present invention respectively. On the chromatic medium plate and the medium plate of the prior art prepared above, turn the plate over and incubate at 35°C for 24h, and do another group at the same time, culture at 30°C for 24h respectively.

2.Aspergillus niger

Aspergillus niger is a mold classified within the Nigri section of the Aspergillus genus. The Aspergillus genus consists of common molds found throughout the environment within soil and water, on vegetation, in fecal matter, on decomposing matter, and suspended in the air. Species within this genus often grow quickly and can sporulate within a few days of germination.

A. niger is a strict aerobe; therefore, it requires oxygen to grow. The fungus can grow in a range of environmental conditions; it can grow at temperatures ranging from 6 to 47°C. As a mesophile, its optimal temperature range is 35-37°C. It can tolerate pH ranging from 1.5 to 9.8. A. niger is xerophilic, meaning it can grow and reproduce in environments with very little water. It can also can also grow in humid conditions even tolerating environments with 90-100% relative humidity. The fungus is most commonly grown on potato dextrose agar (PDA), but it can grow on many different types of growth media including Czapek-Dox agar, lignocellulose agar, and several others.

(1) Take Aspergillus niger (Aspergillus niger) BLCY-02 and inoculate it in a solid medium, and activate it for 20-30 hours under the condition of 30-38°C to obtain an activated strain;

(2) Take the activated bacterial strain obtained in step (1), inoculate it in the seed culture medium, and proliferate and cultivate it for 20-30 hours under the condition of 30-38° C. to obtain the seed liquid;

(3) Take the seed solution prepared in step (2), inoculate it in the fermentation medium at a volume ratio of 1-10%, and expand the cultivation at 30-38° C. for 20-35 hours to obtain the cell fermentation liquid.

Recovery and Passage

●Open the freeze-dried strain tube in an aseptic operation, and add an appropriate amount of culture medium (select the culture medium according to the strain, such as nutrient broth for bacteria, Sandcastle liquid medium for Candida albicans, Sutong for mycobacteria, etc.) Comprehensive liquid medium, Aspergillus niger select malt extract nutrient broth medium), blow and suck several times to melt and disperse the bacteria. Take a test tube containing 5.0 mL ~ 10.0 mL of the corresponding culture solution, drop a little strain suspension, and incubate at a suitable temperature for a specified time (generally 36 ℃ ± 1 ℃ for 18 h ~ 24 h, for Aspergillus niger at 30 ℃ ± 1°C for 42 h to 48 h), this is the first generation culture.

●Use an inoculation loop to take the first-generation culture, or take an appropriate amount of bacterial solution/porcelain beads from the strain freezing tube, and inoculate it on the corresponding medium plate (select the medium according to the strain, such as nutrient agar for bacterial selection). Culture medium, Candida albicans select sandcastle agar medium, mycobacterium selects modified Roche medium or other commercial mycobacteria-specific compound agar medium, Aspergillus niger selects malt extract agar medium), at a suitable temperature Cultivate until the specified time (generally 18 h to 24 h at 36 °C ± 1 °C, 72 h at 36 °C ± 1 °C for mycobacteria, 42 h ~ 48 h at 30 °C ± 1 °C for Aspergillus niger), this is the first time.

2 generation culture.

●Pick typical colonies from the second-generation culture and inoculate them on the corresponding medium slant or plate (select the slant of the medium according to the strain, such as the slant of nutrient agar for bacteria, the slant of sandcastle agar for Candida albicans, the slant of sandcastle agar for mycobacterium Select modified Roche’s medium or other commercial mycobacteria-specific composite agar slant, Aspergillus niger selective malt extract agar medium plate), and cultivate at a suitable temperature for a specified time (generally 36 °C ± 1 °C for 18 h ~ 24 h, mycobacteria at 36 ℃ ± 1 ℃ for 72 h, Aspergillus niger at 30 ℃ ± 1 ℃ for 42 h to 48 h), this is the third generation culture.

●Take the third-generation culture, inoculate it on the slant of the corresponding medium, and cultivate it at a suitable temperature for the specified time. This is the fourth-generation culture.

●Cultivate to the required number of passages according to the above method, and mark the basic information such as the name of the strain, the number of the strain, the number of generations, and the date of passage on the test tube when passaging.

Storage

1)Storage of strains in cryopreservation tubes

Bacteria can be frozen in glycerin, liquid paraffin, porcelain beads, etc., and the appropriate freezing method should be selected according to its characteristics. The cryopreservation tube shall indicate the basic information such as the name of the strain, the number of the strain, and the date of freezing. Bacteria cryopreservation tubes should not be stored at -80 ℃ for more than 18 months. After a certain period of time, the strain cryopreservation tubes can be recovered and frozen again.

2)Vacuum freeze-dried storage

Vacuum freezing technology is used to freeze-dry the bacteria into bacterial powder and store them for a long time. The strain tube should be marked with basic information such as the strain name, strain number, date, etc., and stored at -20°C±2°C

LIB Environmental Chambers for Biology Lab

LIB has been concentrating on theenvironmental test chamber industry for many years, providing high-quality equipment and professional solutions for companies, schools, laboratories, and service centers, helping various organizations to successfully complete many tests, which greatly saves the cost of the enterprise and improves product reliability and durability.

On the other hand, the LIB environmental test chamber has also contributed its own strength to laboratories and various scientific research institutions, and promoted the development of academics.

In the field of biology, the LIB environmental test chamber has unlimited potential and possibilities. It is a medical and biological laboratory equipment used for microorganisms, bacteria, and cell culture. There are many types of incubators. LIB keeps innovating technology and doing its best in every aspect. This article introduces the living and cultivation environment of microorganisms, bacteria and fungi in the laboratory. The above indicators: temperature, humidity, pressure, gas, gas concentration, salt spray environment, light, high and low temperature, constant temperature, etc., can be realized by LIB environmental test chamber.

We provide high-quality and reliable equipment for biological laboratories, making experimental results and data more accurate, and promoting better development in the field of scientific research. If you are interested in this kind of test chamber, please contact LIB immediately.