Chapter-Microbial cell culture and its applications

c) Nitrosoguanidine is used as chemical mutagen, whereas UV rays

a) Recombinant DNA technology is used for strain improvement.

c) Glycerol is used as cryoprotectant as it avoid the formation of ice crystal in liquid nitrogen.

a) Bacteria grow by binary fission.

c) Baffles are used for agitation in a fermenter.

a) Olive oil is used as antifoaming agent in a fermenter.

(b) IMTECH (India at Institute of Microbial Technology is in Chandigarh.

b) The culture preservation at -196° is called as Cryopreservation.

c) The efficiency of chemical reaction is 80- 90% but the microbial conversion efficiency is 20- 30% as the organism for growth and multiplication utilizes a large part of the substrates.

d) Mannitol salt agar is a synthetic media.

a) Cassava is used a source of carbon in a growth media for microbes

Corn steep liquor is a rich source of nitrogen.

a) Sterilization can be achieved by high pressure and heat.

c) Sterilization of nutrient media can be done at 15 psi for 15 minutes.

a) Animal cell culture is sterilized by filtration as such media are completely soluble and contain heat labile components.

c) Bacteria grow at Neutral pH.

a) On large-scale, fermenter is sterilized by steam.

b) When microbes are cultivated in the laboratory aeration and mixing can be done by putting the flasks on the shakers.

a) Impeller is a kind of agitator.

c) In stirred tank fermenter agitation and aeration is done by impeller.

d) After inoculation in fermentor no cell growth occurs for sometimes it is called time of adaptation or lag phase.

a) The cell grows gradually and thereafter show maximum growth rate. This period is called trophophase or log phase or exponential growth phase.

c) In continuous culture the growth retards due to depletion of nutrients not due to accumulation of toxic products.

c) Adding of fresh medium to the fermenter and removal of spent medium and microbial biomass from it results in prolonged exponential phase.

b) In turbidostat the concentration of cells in the culture is kept constant by controlling the flow of medium. Such that turbidity of the culture is kept in narrow limits.

c) Fungi divide by chain elongation and branching.

a) Separation of cells from the fermented broth is done by centrifugation or ultrafiltration.

a) Enrichment liquid culture is a technique resulting in an increase in the number of a given organism relative to the numbers of other types of organism. The process involves taking a mixed population and providing conditions suitable for the growth of desired organism.

c) Microarray technique allows robotic automation that permits the screening of large number of microorganisms.

b) Streptomyces viridifaciens that produces chlortetracycline. Ultraviolet treatment is used to mutate it and from which it may be seen that the mutated strain produces more than twice chlortetracycline than the natural variants.

d) Cells are suspended in cryoprotective agents like 10% glycerol and sealed in ampoules and then frozen in liquid nitrogen.

d) A primary metabolite is directly involved in normal growth, development, and reproduction. Primary metabolites like alcohols, acids, and vitamins are microbial products are produced during exponential growth phase.

c) Vaccines for typhoid, tetanus and tuberculosis have been prepared by using whole cells of bacteria.

c) Tempeh is a Javanese food made from fermentation of soybean by Rhizopus oligosporus into a cake form. Tempeh’s fermentation process and its retention of whole bean give it a higher content of protein, dietary fiber and vitamins.

a) A secondary metabolite is not directly involved in those processes, but usually has an important ecological function. Examples include antibiotics and pigments, and toxins. Secondary metabolite are produced during stationary phase of growth.

d) Microbial cells may be used to convert a compound into a structurally related, financially more valueable, compound. Although the production of vinegar is the most well known microbial transformation process.

b) L- Lysine is produced by Corynebacterium glutamicum.

c) There are a number of readily available commercial media such as LB media, LB broth, nutrient media, nutrient broth, mannitol salt agar, and MacConkey agar etc. These media contain complex components such as peptone, beef extract, yeast extract, and casein. Such media are called semi- synthetic media.

a) Barely grains may be partially germinated and heat-treated to give malt, which contains sugar and starch.

b) The most common cause of foaming is due to proteins in the medium, such as corn steep liquor, peanut meal, soybean meal.

b) foaming can cause removal of cells from medium that will lead to autolysis. If uncontrolled, then numerous changes may occur and physical and biological problems may be created.

c) Sulphonates are used as antifoaming agent

a) Bacteria grow at neutral pH whereas fungi and yeast grow at acidic pH. Therefore before autoclaving the medium the pH is balanced according to the need.

c) Antifoaming agents can be sterilized by heat.

c) It is the simplest type of closed culture system in which microorganism grow in a vessel known as fermenter provided with limited amount of medium containing all the nutrient at optimum environmental conditions.

c) Deceleration phase is also known as stationary phase. It is due to consumption of nutrients, which is called substrate limitation.

a) Culturing microbes in the laboratory, in a ordinary flask, is basically a batch culture.

a) In chemostat growth is controlled by the availability of the growth limiting chemical component of the medium.

b)The cell concentration of the unknown microbial sample can be calculated by measuring the absorbance at the same wavelength.

c) Coulter counter is an electronic instrument and is used for direct counting of microbial cells in suspension.

(b)

Explanation: Corn steep liquor is a by-product of starch; primarily used as nitrogen source, contains lactic acid, sugar and polysaccharides.

(c)

Explanation: Inorganic nitrogen may be supplied as ammonia gas, ammonium salts or nitrates.

(c)

Explanation: Agar is a gelatinous substance derived from seaweed; Gelidium amansii.

(b)

Explanation: Phosphate is used as a buffer to minimize pH.

(b)

Explanation: Foaming is a big problem in microbial culture. Alcohols and esters are used as antifoaming agents.

(c)

Explanation: Energy for growth comes either from the oxidation of medium components or light.

(d)

Explanation: Water is the major component of all fermentation media. For laboratory use, single distilled or double distilled water is adequate.

(d)

Explanation: In microbial processes foaming is a problem.

Foaming can cause removal of cells from medium that will lead to autolysis.

(d)

Explanation: It should disperse readily and have fast action on existing foam. It should be active also at low concentration and nontoxic to microorganism.

(d)

Explanation. Sucrose is the most important sugar in plants. It is generally extracted from sugar cane or sugar beet and then purified and crystallized.

(c)

Explanation. Barely grains may be partially germinated and heat-treated to give malt, which contains sugar and starch.

(c)

Explanation. In most microbial processes foaming is a problem. It may be due to a component in the medium or some factor produced by the microorganism. Commonly used antifoams are fatty acids such as olive oil or sunflower oils.

(b)

Explanation. The most common source of energy is carbon source such as carbohydrates, lipids and proteins.

(a)

Explanation. When microbes are cultured in the laboratory, sterilization of the nutrient medium can be done in an autoclave at 15 psi for 15- 20 minutes or heating the medium at 120°C for 15- 20 minutes.

(d)

Explanation. Viruses normally do not follow a regular growth pattern as they grow intracellularly in host cells.

(b)

Explanation. Cell counting is generally carried out in dilute culture as high cell density presents problems in counting.

(a)

Explanation. Coulter counter is an electronic instrument and is used for direct counting of microbial cells in suspension.

(a)

Explanation. Mutant selection is one of the oldest methods of strain improvement.

The strain is exposed to chemicals (e.g. nitrosoguanidine or NTG) or physical (UV rays) mutagens and the mutants having improved chacteristic are selected.

(c)

Explanation. Streptomyces viridifaciens produce chlortetracycline. Ultraviolet treatment is used to mutate it from which it may be seen that the mutated strain produces more than twice amount of chlortetracycline than the natural variants.

(a)

Explanation. Cultures grown on agar slopes may be stored in a refrigerator (5°) or a freezer (-20°) and subcultured around every six months interval.

(b)

Explanation. This process utilizes the long-term preservation of tissues and cells at ultra-low temperature i.e. in liquid nitrogen (-196⁰C) by using cryoprotectants e.g. glycerol, proline, dimethylsulfoxide and mannitol.

(d)

Explanation. Cryopreservation process utilizes the long-term preservation of tissues and cells at ultra-low temperature i.e. in liquid nitrogen (-196⁰C) by using cryoprotectants e.g. glycerol, proline, dimethylsulfoxide and mannitol.

(a)

Explanation. The cryopreservative DMSO is a small molecule, which is soluble in lipids and generally added to minimize injury to cells during freezing and thawing.

(c)

Explanation. Dried soil cultures have been used widely for culture preservation, particularly for sporulating mycelial organisms.

(b)

Explanation. Lyophilization or freeze drying involves the freezing of a culture followed by its drying under vacuum, which results in sublimation of the cell water.

(c)

Explanation. When microbes are cultured in the laboratory, sterilization of the nutrient medium can be done in an autoclave by heating the medium at 120°C and 15 psi for 15- 20 minutes.

(d)

Explanation. Sparging involves passing sterile air or steam through the medium by having a sparger device at the bottom of the fermenter.

(d)

Explanation. In a fermenter, sparger is used to introduce air or gas into the liquid medium.

(a)

Explanation. The pH and temperature of the medium influences the growth of the microorganism. The normal buffer system in tissue culture media is the CO₂ bicarbonate system.

(b)

Explanation. The disturbance of pH to the addition of acid makes pH control very difficult by conventional means. For this purpose, a pH probe always remains in contact with growth medium.

(c)

Explanation: For small- scale laboratory culture, tubes and flasks can be sterilized in a pressure cooker.

(d)

Explanation: The contamination may be avoided by

a) Using a pure inoculum.

b) Sterlizing the medium.

c) Sterilizing the fermenter vessel.

d) Sterilizing all materials added.

e) Maintaining aseptic conditions during the fermentation.

(b)

Explanation: Sterlization of fermenter vessel is normally done by heating the jacket or coils of the fermenter with steam and sparging steam into the vessel through all the entry points.

Steam pressure is held at 15 psi for 20 minutes.

(b)

Explanation: The normal buffer system in tissue culture media is the CO2 bicarbonate system. This is a weak buffering system and can be improved by use of zwitter ion.

(c)

Explanation: Bacteria grow at neutral pH whereas fungi and yeast grow at acidic pH. Therefore before autoclaving the medium the pH is balanced according to the need.

(c)

Explanation: Microorganisms are classified according to the temperature at which they grow optimally. Temperature controls the rate of reaction that influences the growth rate of the organism.

(a)

Explanation: Filtration is used for animal cell culture sterlization as these media are completely soluble and contain heat labile components.

(d)

Explanation: Hot air oven is widely used in laboratories to sterilize glasswares. The glasswares are dry heated at 180^oC for 20 minutes or 190^oC for 12 minutes.

(b)

Explanation: Hot air oven is electrical device used in sterilization of glasswares. It uses dry heat to sterilize articles. Generally it can be operated between 50 to 300^oC.

(d)

Explanation: According to optimum temperatures for the growth of microorganisms, these are divided into following categories–

Psychrophiles: ~15oC

Mesophiles: ~37oC

Thermophiles: ~55oC.

(c)

Explanation: When microbes are cultivated in the laboratory, aeration and putting the flasks on the shakers can do mixing.

(b)

Explanation: Aerating and agitating (mixing) the fermentation broth normally meet the oxygen demand of the fermenter.

(c)

Explanation: In laboratory, microbes are cultivated in test tubes or Erlenmeyer flasks. Such cultures are carried out in 100- 1000 ml volume.

(b)

Explanation: Baffle flask has a V shaped notch in the sides of flask. This improves the growth of the microbes by providing greater agitation of solutions to improve oxygen or gas transfer.

(c)

Explanation: The first step in production of any microbial product is its research in laboratory. Before going to industrial production it is produced in the pilot plant to standardize the conditions.

(c)

Explanation: After log phase the growth decreases it is called deceleration phase. It is due to consumption of nutrients, which is called substrate limitation, which results in stationary phase.

(b)

Explanation: In continuous culture the production is continuous. The growth medium is formed in such a way that one of the nutrients is in limited quantity. Adding of fresh medium just before the nutrient get exhausted prolongs the growth.

(b)

Explanation: In turbidostat the concentration of cells in the culture is kept constant by controlling the flow of medium. Such that turbidity of the culture is kept in narrow limits.

(a)

Explanation: All these microorganisms grow in different ways

a) Bacteria grow by binary fission

b) Yeast divides by budding

c) Fungi divide by chain elongation and branching

d) Viruses normally do not follow a regular growth pattern as they grow intracellularly in host cells.

Plant Cell Culture and Applications