Quantitative RT-PCR is the most common method used for measuring mRNA levels from small numbers of cells. This type of analysis can be difficult when cell samples are limiting. Ambion's Cells-to-cDNA II Kit (patent pending) overcomes this limitation by using an RT-PCR compatible cell lysis buffer, eliminating RNA isolation altogether. The kit produces cDNA from cultured mammalian cells in less than 2 hours. Here Drs Dziennis and Habecker use the Cells-to-cDNA II Kit to perform real-time RT-PCR on small numbers of primary sympathetic neurons.
Dziennis S and Habecker BA (2003) Cytokine suppression of dopamine beta hydroxylase by extracellular signal regulated kinase-dependent and -independent pathways. JBC 278(18):15897904.
As part of a more detailed study,
Dziennis and Habecker used sympathetic neurons to investigate
the cholinergic differentiation factor suppression of
dopamine--hydroxylase (DBH), a norepinephrine synthetic
enzyme. They measured DBH mRNA levels using real-time RT-PCR
to determine if activation of a MAP kinase pathway was crucial
for suppression of noradrenergic function.