TUDelft/11 September 2008

From 2008.igem.org

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September 11th

Transformation of PCR products

Ligation of PCR products of E. coli genes was performed o/n and a direct transformation of the ligation products was performed. 50 ul of competent cells were added to the 15 ul ligation mix transformed o/n. Results will be checked by colony PCR tomorrow.

Colony PCR of 3A Assembly

The transformants from the assembly have grown, indicating the assembly has worked. We did a colony PCR on a couple of the transformants, to check for insert sizes. The gel of the colony PCR is displayed in figure 1.

Figure 1. Colony PCR of the thermosensitive part / luciferase+Terminator 3A assembly, the lanes indicated by arrows were used to continue assembly, or in the case of K115012, to use as a negative control for the experiments.

We suspect the parts which gave multiple bands at very different sizes are parts which have not ligated inserts but are just a couple of backbones ligated together, as this would give primer annealing zones in all kinds of orientations. For each assembly we've PCR'd a construct of the correct size (some extra bands could be a consequence of the use of B0015 as terminator).