Figure 3.

qPCR measurements of retinal nNOS transcripts using an Applied Biosystems TaqMan™ assay. No difference was seen between control and diabetic nNOS mRNA levels. The values are depicted as mean normalized expression (MNE) compared to the 18s rRNA (n=4). The data were analyzed using the qGene template [30], using the following equation:

$$\begin{array}{l}\text{MNE}=[\frac{{(E_{ref})}^{C{T}_{ref}},replicate1}{{(E_{target})}^{C{T}_{target}},replicate1}+\frac{{(E_{ref})}^{C{T}_{ref}},replicate2}{{(E_{target})}^{C{T}_{target}},replicate2}+\frac{{(E_{ref})}^{C{T}_{ref}},replicate3}{{(E_{target})}^{C{T}_{target}},replicate3}+\frac{{(E_{ref})}^{C{T}_{ref}},replicate4}{{(E_{target})}^{C{T}_{target}},replicate4}]\\ \\ 4\end{array}$$

where; E_target, PCR amplification efficiency of the target gene; E_ref, PCR amplification efficiency of the reference gene; CT_target, threshold cycle of the PCR amplification of the target gene; CT_ref, threshold cycle of the PCR amplification of the reference gene. The CT was defined as the cycle at which the fluorescence rises appreciably above the background fluorescence and determined automatically using Applied Biosystems SDS software version 2.3.