Real-time PCR: what relevance to plant studies?

The appearance of genetically modified organisms on the food market a few years ago, and the demand for more precise and reliable techniques to detect foreign (transgenic or pathogenic) DNA in edible plants, have been the driving force for the introduction of real-time PCR techniques in plant research. This was followed by numerous fundamental research applications aiming to study the expression profiles of endogenous genes and multigene families. Since then, the interest in this technique in the plant scientist community has increased exponentially. This review describes the technical features of quantitative real-time PCR that are especially relevant to plant research, and summarizes its present and future applications.

DEAD-box RNA helicases in Arabidopsis thaliana: establishing a link between quantitative expression, gene structure and evolution of a family of genes.

The model genome of Arabidopsis thaliana contains a DEAD-box RNA helicase family (RH) of 58 members, i.e. almost twice as many as in the animal or yeast genomes. Transcript profiling using real-time quantitative polymerase chain reaction (PCR) has been obtained for 20 AtRHs from nine different organs. Two AtRHs exhibited plant-specific profiles associated with photosynthetic and sink organs. The other 18 AtRHs had the same transcript profile, and the levels of transcription of these 'housekeeping'AtRHs were under strict quantitative control over a large range of values. Transcript levels may be very different between the most recently duplicated genes. The master regulatory element in the definition of the transcript level is the simultaneous presence of a TATA-box and an intron in the 5' untranslated region (UTR). There is a positive and highly significant correlation between the size of the 5' UTR intron and the transcription level, as long as a characteristic TATA-box is present. Our work on the housekeeping AtRHs suggests a scenario for the evolution of duplicated genes, leading to both highly and poorly transcribed genes in the same terminal branch of the phylogenetic tree. The general evolutionary drive of the AtRH family, after duplication of a highly transcribed ancestral AtRH, was towards an alteration of the transcriptional activity of the divergent duplicates through successive events of suppression of the TATA-box and/or the 5' UTR intron.