Project progress

Progress of parts

miniprep of ligY colonies. nannodrop: 111 ng/µl and 116 ng/µl.
restriction digest of ligY miniprep
gelelectrophoresis of restricted ligY and compared to ligA. the ligY fragment was larger than ligA which means that the ligation Y fauled. the promotor region probably ligated to RFP which was not restricted out of the vector properly.
gel extraction of plasmid pSB1A2 (nanodrop: 10ng/µl)
a new ligY was performed
control of pcr-product . It was ok.

part	concentration	260/280
COMT1 1	63,5	1,98
COMT1 2	99,4	1,95
COMT1 3	84,5	1,91
COMT2 1	92,1	2,00
COMT2 2	80,4	1,93
COMT2 3	74,2	1,95
COMT3 1	92,0	2,03
COMT3 2	62,2	2,04
COMT3 3	70,5	2,04
COMT4 1	93,4	1,91
COMT4 2	104,4	1,96
COMT4 3	81,9	2,01

Restriction with S/P, SAMS2 P (was already cut with E/X) and EF3 with E/S
Another attempt at electroporating EFT using EFT ligation from last week
Gel electroforesis of 5 μl from the restriction sample (30 μl)
- Nothing in EF lane (no DNA???)
- Promotor showed again two results of 900bp and 2k bp (wut is going on?)
An attempt was made to seperate the restriction enzymes by PCR purification on the remaining restriction sample, however nanodrop showed very confusing results, conclusion: let's not try this again. Next time heat inactivation of restriction enzymes to make sure they are inactive.
Made mother plate of SAMS

miniprep of virA in the uc vector, fragment was digested a the gelelectrophoresis showed a good result
new restriction digest for virB was done
TOPO-cloning for rpoA failed again, so due to deadlines we decided that there was to few time left to do the TOPO-cloning again or with new protocols and to do the mutagenesis also.