VR vector insert purification continued #

Re-ran BglII digest followed by PCR #

Re-ran a BglII digest of vector but digested for two hours in order to access if the subtle differences in height of the bands are due to under digestion or from sequences differences (DNA secondary structures).

Comparing lanes directly shows that patterns look basically the same, adding evidence to the differences in DNA secondary structure argument as it is unlikely digestion was incomplete in the same ways in both experiments.

Based on these results I went forward with agarose gel extraction of the 2hr digested samples.

Agarose gel extraction yields and follow up PCR #

Did standard freeze and squeeze extraction. Then ODed each sample. Total sample volume was around 20 ul.

After extraction I used 1ul of each sample in a 7ul PCR reaction (running out of master mix and primer) then ODed the reaction contents.