Abstract

In general, metabolic engineering creates a way to have microorganisms be manipulated to create molecules and compounds which we want. Things that can be produced include mass chemicals, fine chemicals, fuels, and pharmaceuticals. If metabolic engineering can be integrated for mass production of something like fuels, we as a society could have a mass source of clean energy which would help to make fuels cheaper and slow down climate change and global warming. This form of engineering could pioneer a whole new area of technology and resource production which could shape the future. Besides resource conservation, metabolic engineering can lead to many biological discoveries that would help our overall health as a society. 

What is a Sigma Factor?

Sigma factors are the essential proteins in prokaryotic cells that allow for transcription to occur. Transcription is the process by which DNA is transcribed into messenger RNA (mRNA). Sigma factors direct RNA polymerase to a promoter region, or a specific binding site just upstream of the gene to be transcribed, which serves as the starting point for transcription. This process starts only once the sigma factor guides RNA polymerase into position. In prokaryotic cells, ribosomes translate the mRNA into polypeptides as soon as it is transcribed. These polypeptides are then used to build the protein encoded by the transcribed gene. Sigma factors recognize unique promoter sequences, which allows the cell to regulate gene expression in response to environmental factors like heat or nutrient availability. By making changes to promoter sequences and observing their effects, we can gain insight into the importance of specific DNA sequences required for transcription and how conserved they need to be for successful gene expression

Procedure

We would design the binding site for ECF 11 with the specific changes we wanted and order those DNA sequences. Then we would assemble those DNA sequences into plasmids with the binding sites and the GFP reporter gene using centrifuge. After that we did colony PCR, picked the 3 best colonies and perform gel electrophoresis, looking at how far our plasmids traveled in the gel and compared the length of the plasmid we got from doing gel electrophoresis to the length of the plasmid we wanted to confirm we created the plasmid correctly. We then transformed the plasmid into E. coli using heat shock. Finally, we took all of our created E. coli samples, saw how much the sampled glowed green and measuring the optical density, converting that to fluorescence, and comparing these results with the controls.

Career Skills Gained

• Communication: Learned to communicate with others about what work needed to be done in a professional way

• Critical Thinking: Learned by looking at the past data and understanding it to conduct these new experiments

• Teamwork: Learned by collaborating on the common goals we had by wanting to get good results from our experiments.