Part:BBa_M50447

Title - An attempt to mimic pancreatic beta cells. We connected a glucose sensitive promoter (sourced from iGEM team IGEM14_Tshingua) to a yeast kozak for transcriptional purposes. After the yeast kozak was a single chain insulin gene connected to a his-tag. We connected the his-tag so we could check for protein production with a Western blot. After the his-tag was a t2a peptide so we could attach an EGFP reporter gene. The t2a was intended to allow the ribosome to transcribe two genes given only one promoter. After the EGFP gene was the terminator from the iGEM parts registry (part number BBa_K1462070) with the consensus sequence for a poly A tail to protect the mRNA from degradation after it is exported from the nucleus.

Description - The constituent parts were intended to mimic pancreatic beta cells. Beta cells are what's missing from a person who has type 1 diabetes, and importantly, beta cells turn on insulin production when blood glucose is high. Then, insulin signals to fat, liver, and muscle cells to uptake blood glucose and use it as energy, thereby effectively regulating blood glucose. However, people with type 1 diabetes do not have beta cells so they need to get their insulin delivered in the form of pens, pumps, shots, etc. This was a project trying to use a glucose sensitive promoter to express insulin and EGFP given high concentrations of glucose. It did not work as intended, i.e. we saw no increase in insulin/egfp expression given increase in concentrations.