Difference between revisions of "Part:BBa K3174008"

Revision as of 16:24, 11 October 2019

SYFP2 coding sequence with IS hotspot removed

This is a redesigned version of the Super Yellow Fluorescent Protein 2 (BBa_K864100) that has had a region that was prone to insertion element mutations altered. This removal of a mutation hotspot resulted in the expression of SYFP2 in culture being maintained for a longer period of time than the original plasmid.

Usage and Biology

This part codes for the bright yellow fluorescent protein SYFP2, which has an excitation peak of 515 nm and an emission peak of 527 nm.

Testing and Measurement

Cultures of TOP10 E. coli were transformed with either the original SYFP2 plasmid or the redesign and propogated over four days. Each day a new culture was created using a 1:1000 dilution of the previous day's culture and allowed to grow for 24 hours at 37 C. Fluorescence was measured daily using flow cytometry. The data showed that while the original plasmid stopped expressing fluorescence after one day, the redesigned plasmid retained expression for up to four days.

Figure 1: Fluorescence histograms for TOP10 cells with the original SYFP2 plasmid. The x-axis shows the intensity of fluorescence on a logarithmic scale, and the y-axis displays the number of cells. Each line represents a different day of propagation, and the shaded gray curve is a blank. Peaks to the right of the blank represent fluorescent cells while peaks on or to the left of the blank represent cells that have ceased to be fluorescent. All replicates were grown under the same conditions.

Figure 2: Fluorescence histograms for TOP10 cells with the redesigned SYFP2 plasmid. The x-axis shows the intensity of fluorescence on a logarithmic scale, and the y-axis displays the number of cells. Each line represents a different day of propagation, and the shaded gray curve is a blank. Peaks to the right of the blank represent fluorescent cells while peaks on or to the left of the blank represent cells that have ceased to be fluorescent. All replicates were grown under the same conditions. No data could be collected for Day 3 of Replicate 4.

Sequence and Features