Restriction SAMS1 1 X/P, S/P, EF1 and EF2 E/S, COMT4 2 and COMT1 1 E/X
SAMS and EF show expected results, promotor shows the same result as yesterday (1 part of 900bp and 1 of 2k bp) and COMT looks good.
Ligation of EF + TER, EF was not purified from gel, instead the enzymes were heat inactivated on 65C for 15 min, TER was an older vector which was purified
Promotor transfered from glycerol (-80) to a plate and liquid culture.
At this point we realised the promotor was not in a standard vector but rather in which contains an additional RFP after the Spe restriction site, thus a part of 900bp and one of 2k bp is expect when digesting with P
Because virA was now properly inserted in the pucvector we could now start mutagenesis on restriction sites 187 and 720. We did this with PCR. We used a protocol for site-directed-mutagenesis with the matching primers. PCR was done overnight
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