BIOTEC Dresden/Notepad/9 September 2010

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Revision as of 12:15, 11 October 2010

Parts Assembly

Eight colonies were picked for each part and colony PCR was done.

4a, 9b, 14f, 20f, 21f

The PCR samples were run on an agarose gel and the bands obtained did not correspond to the desired insert length. This suggests that the ligation did not work and hence, the primers alone were amplified without the desired part amplification.

The digest of the plasmid backbone might not have occurred as desired and hence the backbone was again digested and purified by PCR purification. The concentration of the backbone was measured. A 1:3 ligation was done for the following parts.

4a, 9b, 14f, 20f, 21f

The ligation mix was transformed by electroporation and plated and left for overnight incubation for growth of colonies.

Fusion Protein

A colony PCR was performed to confirm the insertion of our fused insert into the plasmid pETMM43 or pETMMZZ. 15 clones from each plate were picked and reaction conditions optimized. No positive clones for the aiiA construct were found.