May 05th, 2010
Today I made a PCR of the fragment of 100bp obtained from the restriction with EcoRI and PstI of plasmid pSB1AK3. I prepared 4 different samples each obtained from a different colony.
This PCR was made following the same protocol from the last time with a few changes: I put 1μL of template DNA without any dilution and 17.5μL of MgCl2 (to avoid the quenching of all the Mg+ by the excess of DNA).
I just used a positive control; a PCR mix that only lack Taq Polymerase.
When the PCR was accomplished I made an agarose gel of 1.5% to prove my results.
At least the PCR was successful; I obtained 4 lines of 100bp corresponding to the fragment containing the double terminator. We get a double terminator (BBa_B0015).
However we also obtained some smaller bands and we don't know what are them; they could be due to inespecific primer annealing.