Week of 9/23/24 Double Digestion

In order to assemble the PgK (PgRNA and Kanamycin) fragment onto the guide plasmid, we still needed to have successful double digestion of PgRNA with enzymes Xbai and BAM. BAM has worked without failure, but we are inhibited in our progress due to the Xbai fragment not ligating despite the primers being coded specifically for this fragment. We are not sure why this may be, so we are narrowing down on what exactly is the problem whether or not the primers are bad, if the enzyme itself is bad, or if the double digestion is not suitable for Xbai to ligate. Going forward, we will only single digest Xbai and find a basis for which the issue is. We have also brainstormed a name for our plasmid, PVELM (Plasmid and the first initial of the prime researchers within our group that have been working on this since summer, Vanshika, Emme, Lani, Madi). 

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