Application of real-time RT-PCR quantification to evaluate differential expression of Arabidopsis Aux/IAA genes

The molecular techniques including Northern blot, dot blot, in situ hybridi zation, etc. have been successfully used to estimate semi-quantitatively mR NA levels in plant samples. In this study, we employed a real-time reverse transcription-PCR (RT-PCR) assay using SYBR Green I fluorescence methodolog y to evaluate accurate quantitation and sequence specific detection of Aux/ IAA mRNA levels in Arabidopsis. Results obtained indicate a linear dynamic range of 10(2)-10(6) Aux/IAA mRNA copies with standard deviations of genera lly less than 15%. As a model experiment, the outcome of analysis of expres sion patterns of five Aux/IAA genes in Arabidopsis under various chemical a nd temperature treatments is presented. The method presented here provides a sensitive and rapid technique to evaluate plant Aux/IAA mRNA expression l evels in nanogram order.