Screening Rnai Transformants Of Chlamydomonas For Reduced Expression Of The Photoreceptor Cryptochrome

Jeremy Webb, Western Kentucky University

Abstract

Chlamydomonas reinhardtii is a unicellular green alga. In C. reinhardtii, three types of photoreceptors are known to be present: rhodopsins, phototropins, and cryptochromes. The single cryptochrome is the most likely photoreceptor for adjusting the circadian clock to the daily light/dark cycles, because cryptochromes are involved in clock entrainment in higher plants and insects. In this segment of the research, C. reinhardtii strains, which were genetically modified through transformation with a RNA interference construct, were screened for reduction in cryptochrome compared to the control strain. After C. reinhardtii cultures were harvested in complete darkness, all soluble proteins were extracted from the cells. The proteins were subjected to Western blot (immunoblot) analysis with chemiluminescent detection. Quantification of the images obtained with a digital camera revealed cryptochrome reductions between 0 and 50 percent in the ten strains tested. Should the strains with 50 percent reduction show a reduced ability to entrain to light/dark cycles, such results would suggest that cryptochrome is the primary photoreceptor that controls circadian clock resetting in C. reinhardtii.