IGEM:Hong Kong HKUST/Investigations/Effect of overloading cells on growth rate/Entry Base: Difference between revisions
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==Authors== | ==Authors== | ||
*[[User:Chloe T.C. Tang | TANG, Chloe T.C.]] | *[[User:Chloe T.C. Tang | TANG, Chloe T.C.]] | ||
* Tam, L.F. | * Tam, L.F. | ||
* Tam, K.M. | * Tam, Sabrina K.M. | ||
* Tang, Mandy L.Y. | * Tang, Mandy L.Y. | ||
* Siu, Mona M.Y. | * Siu, Mona M.Y. | ||
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==Abstract== | ==Abstract== | ||
It is hypothesized that strong gene expression will affect the growth and viability of cells. This experiment was conducted by incorporating a strong promoter from the Anderson Family (BBa_J23101: Registry of Standard Biological Parts) with a | It is hypothesized that strong gene expression will affect the growth and viability of cells. This experiment was conducted by incorporating a strong promoter from the Anderson Family (BBa_J23101: Registry of Standard Biological Parts) with a strong ribosome binding site (BBa_I13003: Registry of Standard Biological Parts) in <em>Escherichia .coli</em> (<em>E.coli</em>) to test the hypothesis. Quantitative results were obtained by optical density using a spectrophotometer with UV light of 600nm. The results indicated that the effect of overloading cells on growth rate was insignificant. | ||
==Introduction== | ==Introduction== | ||
Gene expression in bacteria involves the production of Messenger RNA (mRNA) from DNA template and translation of the mRNA into protein. These two processes are driven by the binding of RNA polymerase to the promoter region of the gene and the binding of Transfer RNA (tRNA) to the start codon nearest to the ribosome binding site (RBS) respectively. Therefore, to strengthen the expression of a foreign gene in ''E. coil'', strong promoters and RBS are selected as two controlling factors. | |||
Under the stress of being transcribed too often, much energy and materials are required for gene expression. It is suspected that DNA overloading will lower cell viability as energy and materials are wasted during gene expression. Also, the accumulation of proteins may perturb cellular function which in turn affects normal cell growth.Therefore, we would like to investigate the effect of overloading on cell growth and viability. | |||
==Methods and Materials== | ==Methods and Materials== | ||
To investigate the effect of strong gene expression on cell growth and viability, a gene encodes for yellow fluorescent proteins with strong promoter and RBS were constructed and transformed into ''E. coil''. Then, the growth rates of cells with strong promoter and RSB are compared with those without to test the hypothesis. | |||
==Results and Interpretations== | ==Results and Interpretations== | ||
[[Image:OD 600 project.jpeg|850px|center]]<br><br> | |||
As shown in <strong> Figure 1, </strong>the DNA samples for both the experiments with strong gene expression and the negative control show similar growth rate. The results indicated that the effect of overloading cells on growth rate was insignificant. However, all the samples showed an unexpected growth pattern as they did not have a sign of reaching to a steady state. Also, after approximately 235 minutes, the growth rate of all the DNA samples suddenly increased and were not consistent with the previous rates. | |||
==Discussion== | |||
According to J. Monod ("The Growth of Bacterial Cultures",1949), a normal cell growth pattern, in theory, should shows a rapid growth at the beginning, then slows down to reach a steady state and die off at the end. However, all the experimental samples showed an unexpected growth pattern as they did not have a sign of reaching to a steady state. It is suggested that the number of measurements taken might not be enough for showing such a pattern. Also, after approximately 235 minutes, the growth rate of all the DNA samples suddenly increased and were not consistent with the previous rates. | |||
In our hypothesis, it is expected that strong gene expression will affect the growth and viability of cells. However, our results show no differences between the experimental setup with strong gene expression and the control in terms of growth rate. One possible explanation is that the promoter (BBa_J23101: Registry of Standard Biological Parts) and ribosome binding site (BBa_B0032: Registry of Standard Biological Parts) that we used in our experiments might not be strong enough and overloading of cells were not generated. Therefore, their growth rates show similar pattern. Another possible reason is that overloading of cells might not have significant effects of its growth rate. Hence, it is hard to make concrete conclusion simply base on the results obtained. Further investigations are needed to test the hypothesis. | |||
==Conclusion== | ==Conclusion== | ||
Both the experimental cells and the negative control showed a similar growth pattern and rate. It is concluded that cells with our synthesized promoter and ribosome binding site have insignificant effect on its growth rate. | |||
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Revision as of 08:12, 28 January 2015
 Effect of overloading cells on growth rate
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