Stationary phase cell growth

For the identical experimental conditions, electroporation efficiency depends on the type of cells the composition of the membrane, shape, and size of cells strongly influences the electroporation efficiency [40-42]. In electroporation of bacteria, the growth phase of cell has significant influence on transformation efficiency, which is higher for cells harvested and electroporated from mid-log phase. However, cells from stationary phase can also be transected with reasonably good efficiency. Mammalian cell can be electroporated at relatively lower fields but pulse length controls the entry of external molecules into cells. 

Fig. 6. Variation of the levels of shikimate dehydrogenase in N. silvestris during the various growth phases following subculture from cells in stationary phase. The inset shows the constant and reduced levels of activity obtained from cultures maintained continuously in exponential phase (data shown for the 8-to-ll generation interval).

Consider again a closed volume with uniform composition and temperature — in practice a batch reactor, in which only changes with time occur. From its initial composition the medium evolves after seeding by an inoculum, which is a small amount of the living cell culture with optimized composition, through respectively a lag phase, an exponential growth of the number of cells, a stationary phase during which the increase in number of cells is compensated for by their destruction and a death phase with an (exponential) decrease in the number of cells. 

Fig. 1.18. Yeast growth and survival curves in a grape juice medium containing killer toxin (Barre, 1992) +, 10% K2 strain active culture supernatant O, 10% supernatant inactivated by heat treatment, (a) White juice, pH 3.4 cells in exponential phase introduced at time = 0. (b) Same juice, cells in stationary phase introduced at time = 0. (c) Red juice extracted by heated maceration, pH 3.4 cells in exponential phase introduced at time = 0...

Batch fermentation is the most widely used method of amino add production. Here the fermentation is a dosed culture system which contains an initial, limited amount of nutrient. After the seed inoculum has been introduced the cells start to grow at the expense of the nutrients that are available. A short adaptation time is usually necessary (lag phase) before cells enter the logarithmic growth phase (exponential phase). Nutrients soon become limited and they enter the stationary phase in which growth has (almost) ceased. In amino add fermentations, production of the amino add normally starts in the early logarithmic phase and continues through the stationary phase. 

Once there is an appreciable amount of cells and they are growing very rapidly, the cell number exponentially increases. The optical cell density of a culture can then be easily detected that phase is known as the exponential growth phase. The rate of cell synthesis sharply increases the linear increase is shown in the semi-log graph with a constant slope representing a constant rate of cell population. At this stage carbon sources are utilised and products are formed. Finally, rapid utilisation of substrate and accumulation of products may lead to stationary phase where the cell density remains constant. In this phase, cell may start to die as the cell growth rate balances the death rate. It is well known that the biocatalytic activities of the cell may gradually decrease as they age, and finally autolysis may take place. The dead cells and cell metabolites in the fermentation broth may create... 

In phase B it is assumed that the inoculum has adapted itself to the new environment and growth then proceeds, each cell dividing into two. Cell division by binary fission may take place every 15-20 minutes and the increase in numbers is exponential or logarithmic, hence the name log phase. Phase C, the stationary phase, is thought to occur as a result of the exhaustion of essential nutrients and possibly the accumulation... 

The distribution of PG (PG) as culture medium supernatant, cell-wall associated and cell-bound enzyme was observed in K. marxianus during the time course of growth in 5% glucose medium (Table 2). PG secretion started between 8 and 12 h after inoculation and approximately 90% of total PG was secreted in early stationary phase. PG was not detected intracellularly after 24 h of growth. 

Sub-cellular fractionation of five strains reveal the same numbers of bands. The distribution of PG activity in sub-cellular organelles was broadly similar in these five strains. PG activity was detected in low-density vesicles, vacuoles and ER fractions in samples harvested during the early exponential phase of growth. However, PG levels were always lower (at least 1.5 fold) than those found in wild type. Cells of the mutants harvested during stationary phase of growth showed that 84% of total intracellular PG activity was located in the vesicle fraction. No intracellular PG activity was found in stationary phase wild type cells. 

Following the stationary phase the rate of cell death exceeds that of cell growth, and the cell number begins to decrease, resulting in the final death rate part of the curve. 

The development of large-scale cultures involved transferring cells from stock cultures to a series of two liter Fernbach flasks containing enriched seawater medium. After the early stationary phase of growth had been reached (approximately 15-20 days) each of these cultures were used to innoculate 18 liters of the same medium in 20 liter carboys. Large-scale cultures were grown under continuous light (4300 lux cool white fluorescent) at 27.0° C. 

Cells were harvested by centrifugation after cultures reached the early stationary phase of growth (30-35 days). 

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