Stationary phase, inclusion body

In each culture the fraction of cells containing visible inclusion reached an approximately constant value during exponential phase growth (Figure 4). As the cells reached late exponential or early stationary phase the fraction then rose sharply so that eventually 90-95% of the cells contained at least one inclusion body. 

As the cell density increased to 2 x 10 CFU/mL there was a rapid increase in the fraction of cells containing inclusion bodies in all cultures. Regardless of inoculum density, all cultures reached about 90-95% of the cells containing at least one inclusion in stationary phase. 

The densitometer measurements show that the fraction of cells containing visible inclusion bodies correlated to the (relative) amount of prochymosin in the culture as a whole (Figure 7). This correlation appeared to hold both within a single culture as it entered stationary phase and between different cultures with different inoculum sizes. The correlation was fairly linear until about 80% of the cells contained inclusion bodies then the amount of prochymosin sharply increased relative to the other protein band. This corresponded to early stationary phase when there was a sudden increase in prochymosin cortent and an appearance of multiple inclusion bodies within each cell. 

Serial liquid-to-liquid subculture experiments were run to determine the reason for the rapid takeover by non-inclusion body-forming cells. When subcultures were taken from stationary phase cells, the fraction of inclusion body containing cells in the population in stationary phase dropped dramatically to essentially zero within two subcultures. When subcultures were obtained from exponentially growing cells, the fraction of cells showing inclusion bodies in stationary phase dropped much more slowly with each subculture, 84% of the stationary phase cells showed visible inclusions even after four subcultures. 

AU the cultures, regardless of inoculum size, resulted in at least 90% of the ceUs showing at least one inclusion body in stationary phase. 

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