Bacterial Terminator Efficiency and Expression of Two Reporter Genes

Abstract

Terminator sequences regulate gene expression and contribute to genetic diseases and cancer development. In an effort to understand the mechanisms and efficiency of terminators, we characterized bacterial terminators in three synthetic biology constructs. Our test construct encoded a constitutive promoter, green fluorescent protein (GFP), terminator, and pink chromoprotein. Control 1 encoded the promoter, GFP, and terminator. Control 2 encoded the promoter, GFP, and pink chromo. A fluorometer was used to measure GFP fluorescence, and all constructs exhibited high-levels of fluorescence. We used a spectrophotomer and visible appearance to compare the expression of pink chromoprotein in all constructs and investigate the impact of terminators on gene expression. Control 1 did not show evidence of pink chromoprotein expression. Control 2 exhibited absorbance in the pink spectra and was visibly pink. The test construct showed minimal pink spectra absorbance and did not appear pink. The ability of Control 2 to exhibit both fluorescence and pink coloration suggests two reporter genes can be produced via the same promoter. The lack of pink coloration in our test construct demonstrates that the terminator can block the visual appearance of pink chromoprotein but is not strong enough to inhibit low levels of expression detected by the spectrophotometer.