Team:WPI-Worcester/Harvard-Collaboration
WPI and Harvard Biodesign Collaboration
WPI
We completed the biofilm assay linked above with strains of E. coli with modified pili created by the Harvard team. The strains we tested were JW4283, a fim H knockout and negative control for biofilm formation, and C2M+C5 and C9-3+C5, which express modified pili when induced by arabinose and rhamnose. Upon receiving agar stabs from the Harvard team, we streaked plates and incubated them at 37oC for 24 hours. Afterwards, we placed the plates in the refrigerator until we received the arabinose and rhamnose needed to induce pili formation in the experimental strains. Once we had all necessary materials, we grew 5mL liquid cultures of each of the 3 strains in LB in accordance with the protocol. The JW4283 culture was not supplemented with antibiotics or inducers, but the C2M+C5 and C9-3+C5 cultures were supplemented with 5 μL of chloramphenicol, 5 μL of ampicillin, 0.01% arabinose, and 0.5% rhamnose. We prepared the 1:100 dilutions in LB broth and M9 minimal media, which we found to support biofilm formation while designing our biofilm assay. The remainder of the protocol was left unchanged. Below are the results of the assay. A photo of the biofilm plate after it was stained with crystal violet on 9/3/15. Although there appears to be some remaining crystal violet, biofilm formation was not robust in any of the strains.
A table of the OD595nm for each well with the blank subtracted. Average ODs for each strain in both LB and M9 are also included.
A graph of average OD595nm for C2M+C5 and C9-3+C5 in M9 minimal media standardized to average OD595nm for JW4283 in M9 minimal media (indicated by the line at 1). Error bars=+/- standard error of the 4 ODs measured for the two strains in M9.
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According to the results we collected, the Harvard strains with modified pili do not form biofilms under the conditions we subjected them to. These strains may be able to form biofilms under different conditions. Further investigation would have to be done to determine if different if different media types, incubation periods, or transformation into different strains of E. coli could induce biofilm formation. Determining optimal biofilm formation conditions could allow the Harvard team to use the biofilm assay to gather quantitative results for their various binding assays. However, it is possible that the modified pili interfere with biofilm formation. In that case, the biofilm assay would not be a valid method of quantification. Harvard
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