Real-time Quantitative RT-PCR method for estimation of mRNA level of CCAAT/enhancer binding protein in the central nervous system of <i>Lymnaea stagnalis</i>
2004
Hatakeyama, D. | Sadamoto, Hisayo | Ito, E.
The fluorescence-based real-time reverse transcription polymerase chain reaction (RT-PCR) is becoming widely used to quantify mRNA level in cells and tissues and is now a crucial tool for basic biological researches and biotechnology. In the present study, on the basis of the real-time quantitative RT-RCR, we detected and quantified mRNA copies of the transcription factor, CCAAT/enhancer binding protein (C/EBP; an immediate-early gene that is involved in synaptic plasticity and learning and memory) in the central nervous system of the pond snail <i>Lymnaea stagnalis</i>. We designed the primer set and the probe in the specific insert for the detection of <i>Lymnaea</i> C/EBP (LymC/EBP) clone 1. This insert is not contained in LymC/EBP clone 2 by alternative splicing. The copy number of LymC/EBP clone 1 was linearly decreased relative to the dilution of cDNA, and it was estimated 30 copies/ml in test sample. The availability of the present study showed that the real-time quantitative RT-PCR technique is more accurate and more specific for the detection and quantification of the mRNA level of genes in <i>L. stagnalis</i> than the other PCR methods.
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