"
Team:Nanjing-China/pre
From 2013.igem.org
Revision as of 15:09, 27 September 2013 by GaoJian NJU (Talk | contribs)
- Pre-experiments
-
Through consulting literatures and materials, we found that the concentration of atrazine used in experiments related to its degradation was considerably changeable. As we all know, atrazine, which is toxic to bacteria, can reduce the amount of bacteria and influence the efficiency of degradation. To avoid this problem as well as to provide the public with a standard of atrazine concentration in experiments, we conducted a series of pre-experiments, studying the relationship between atrazine and bacteria.
Our primary chassis for wet lab is called K-12, a strain isolated from E. coli. It's a primary model organism with clear genetic background and handy genetic modification. This strain has been widely distributed to laboratories across the world[1]. In addition, chemotaxis in E. coli is well documented[2] so that we can control the movement of it by engineering the proteins in the pathway. Most importantly, the strain E. coli K-12 is a debilitated strain which does not normally colonize the human intestine, and it is not known to have adverse effects on microorganisms or plants.[3]
Experiments and Results
The bacteria strain we used is called K-12, a strain isolated from Escherichia coli. It is a type strain which moves the fastest in soil of all strains with clear genetic background and handy genetic modification.
As Fig. 3-2-1 shows, when the concentration of atrazine was between 200μM and 500μM, the trend of the growth curve was similar to the natural growth curve without any atrazine. However, when bacteria were exposed to atrazine of 1000μM, the growth curve of K-12 changed a lot, especially in plateau period.
Fig. 3-2-1Growth Curve of K-12 cultured in different concentration of atrazine. Atrazine could not restrain bacteria growth significantly if the concentration was between 200μM and 500μM, but the growth curve changed a lot when the concentration of atrazine rose to 1000μM, suggesting that high concentration of atrazine is harmful to K-12.
In order to make their relationship more intuitive, we performed another experiment, comparing the amount of bacteria in plateau period. As illustrated in Fig. 3-2-2, the amount of bacteria cultured in atrazine of 200μM or 500μM hardly changed, while it declined sharply when cultured in concentration of 1000μM.
Fig.3-2-2The amount of bacteria cultured for 10h in different concentration of atrazine. After 10h, plateau period had been reached. It was obvious that the maximum of bacteria exposed to atrazine of 200μM or 500μM hardly changed, but it became much lower when the concentration turned to 1000μM.
Taking the condition of bacteria's growth for consideration, we finally chose 200μM-500μM as the concentration of atrazine in our all experiments.
Reference
[1] Riley M, et al. Escherichia coli K-12: a cooperatively developed annotation snapshot-2005. Nucleic acids research, 2006, 34 (1): 1-9.
[2] Chelsky D and Dahlquist FW. Chemotaxis in Escherichia coli: associations of protein components. Biochemistry, 1980, 19 (20): 4633-4639.
[3] Escherichia coli K-12 Derivatives Final Risk Assessment. (Online) Available from: http://epa.gov/oppt/biotech/pubs/fra/fra004.htm
