Figure 7 shows the performance of our self-designed vector with the DiaMIX and the commercial mix. As can be observed, the DiaMIX still performs better than the commercial kit. However, the difference is not as obvious as before. This indicates that the DNA does not yet posses an optimal structure for an effective performance in a cell-free system. This could be due to differences in the 5' upstream sequence which was demonstrated to have great influence on the initiation of transcription. Furthermore, tags in the beginning of the coding sequence also enhance the expression level as shown by Haberstock, S. et al. 2)
Our self-produced DiaMIX performed about as good as the commercial kit as it is shown in figure 7. Compared to the background, an expression time of two hours resulted in a 2-fold increase of relative fluorescence in both systems. The background fluorescence was estimated by the negative control.