Part:BBa_B0012:Experience

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••••• iGEM Athens 2019

We attempted to measure the efficiency of the terminator by cloning it into the pSB1A10 plasmid and culturing the bacteria for 24 hours (37 degrees Celsius, 200RPM, 0.1% arabinose, 100 µg/mL ampicillin). The culture was pelleted and the pellet was resuspended in 1mL of 1X PBS. From that, 10 μL were diluted in 990 mL of 1X PBS to yield approximately 5 million cells. For FACS, we used BD FACSCanto™ II. During FACS, a low flow rate was used. We used the following excitation and emission wavelengths: GFP: excitation = 488 nm, emission = 510 nm ; RFP: excitation = 488 nm, emission = 584 nm. FACS measurement. The terminator was named T7T for our project. The efficiency is measured through the following formula: Termination efficiency = 1 -{[RFPterm/GFPterm]/[(RFPcontrol/GFPcontrol)mean]}. This measurement showed that the efficiency of the terminator is 74.2%. We also measured the terminator efficiency using the GFP+RFP+ population and the formula: Terminator efficiency = 1 - (Intensity of GFP+RFP+term/Intensity of GFP+RFP+control). This measurement showed that the terminator efficiency of the terminator is 92%.

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••••• KAIT_JAPAN

We have succeeded to confirm this parts.If you want to see the result,go to our wiki(http://2015.igem.org/Team:KAIT_Japan/contribution)

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