Semi-quantitative RT-PCR analysis of photoregulated gene expression in marine diatoms

The low cell densities of diatoms and other phytoplankton in culture has pr ecluded the use of classical RNA analysis techniques for routine studies of gene expression in large numbers of samples. This has seriously hampered s tudies of the basic biology of such organisms. To circumvent this problem, we have developed a high-throughput semi-quantitative RT-PCR-based protocol and used it to monitor expression of a gene encoding a fucoxanthin, chloro phyll a/c-binding protein (FCP) in the centric planktonic diatom Thalassios ira weissflogii. Analysis of FCP gene expression in dark-adapted diatom cul tures revealed that mRNA levels increase 5- to 6-fold in response to white light irradiation and peak around 6 to 8 h. To determine the photoreceptors involved in this response action spectra of FCP gene expression were deter mined using the Okazaki large spectrograph. Responses consistent with the p resence of cryptochrome-, rhodopsin- and phytochrome-type receptors could b e detected. The apparent presence of phytochrome-mediated responses is of p articular interest given the low fluences of red and far-red light waveleng ths in the marine environment.