Difference between revisions of "Part:BBa K1033916:Experience"

Hong Kong-CUHK iGEM 2017

Fluorescent properties of amajLime

Instead of chromorptein, we proposed that amajLime is a fluorescent protein with the property of showing pigmentation, like BBa_E1010. We charaterised spectral properties of amajLime and found its max excitation and emission wavelength at 445 nm nd 485 nm respectively.

Fig.1 Emission and Excitation spectra

Charaterization of pH stability of amajLime

We transformed part BBa_K1033916 with constituitive promoter: J23100 in C41 and grew in 2XYT for 24 hours. Purifying the amajLime by Ion Exchange Chromatography and Hydrophobic Interaction Chromatography, we measured the fluoresece of purified amajLime, which is diluted to 10µg/100µl (total 200µl) in triplicates, in different buffers (ranges from pH2 to pH12). The result shows that the stability drops dramatically in condition below 6 and attains maxima fluorescence at pH 8.

Fig.2 Vary pH attributed to different fluorescent intensity of RFP

Table 1 Plate reader setting of fluorescent measurement

Basic Settings
Measurement Type	Fluorescence
Microplate name	COSTAR 96
Scan mode	orbital averaging
Scan diameter [nm]	3
Excitation	470-15
Emission	515-20
Dichronic filter	auto 491.2
Gain	500
Focal height [nm]	9