The molecular techniques including Northern blot, dot blot, in situ hybridization, etc. have been successfully used to estimate semi-quantitatively mRNA levels in plant samples. In this study, we employed a real-time ...The molecular techniques including Northern blot, dot blot, in situ hybridization, etc. have been successfully used to estimate semi-quantitatively mRNA levels in plant samples. In this study, we employed a real-time reverse transcription-PCR (RT-PCR) assay using SYBR Green I fluorescence methodology to evaluate accurate quantitation and sequence specific detection of Aux/IAA mRNA levels in Arabidopsis. Results obtained indicate a linear dynamic range of 102-106 Aux/IAA mRNA copies with standard deviations of generally less than 15%. As a model experiment, the outcome of analysis of expression patterns of five Aux/IAA genes in Arabidopsis under various chemical and temperature treatments is presented. The method presented here provides a sensitive and rapid technique to evaluate plant Aux/IAA mRNA expression levels in nanogram order.展开更多
基金J. Liu was supported by the JAERI Research Fellow Program.
文摘The molecular techniques including Northern blot, dot blot, in situ hybridization, etc. have been successfully used to estimate semi-quantitatively mRNA levels in plant samples. In this study, we employed a real-time reverse transcription-PCR (RT-PCR) assay using SYBR Green I fluorescence methodology to evaluate accurate quantitation and sequence specific detection of Aux/IAA mRNA levels in Arabidopsis. Results obtained indicate a linear dynamic range of 102-106 Aux/IAA mRNA copies with standard deviations of generally less than 15%. As a model experiment, the outcome of analysis of expression patterns of five Aux/IAA genes in Arabidopsis under various chemical and temperature treatments is presented. The method presented here provides a sensitive and rapid technique to evaluate plant Aux/IAA mRNA expression levels in nanogram order.
