User:Anthony Salvagno/Notebook/Research/2009/03/13/Digestion, Ligation, Transformation Retry Day 2

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I completed the digestion. I had some unsettling results, so we are going to run a gel to see what's going on.

Nanodrop readings: pRS DNA - 714ng/uL, gDNA - 245.6ng/ul

Those number are bad because we had 5uL of DNA (each) and in 25uL of buffer, the above results are way more than that.

Estimations

Kelly estimates that there are about 100ng/uL in each tube (each kind of DNA). I don't know anything about this, but I have a feeling that that is a big overestimation. We'll see. The reason I say that is because last time we did a EtOH precipitation, I got less than 50% yield. Why would I have more this time? Kelly should get a better estimation when the gel is finished.

Results

Apparently I run a beautiful gel. I got good band spacing and nice bright bands. The plasmid DNA had a good cut (no other structures), but the XhoI was mostly undigested. We decided to clean the gDNA with another P/C extraction and EtOH precipitation. Hopefully on Monday I will get some good cutting with XhoI.

As far as the estimations are going, Kelly estimated almost 100% yield from the extraction from the other day. I'm skeptical, but what do I know? Maybe I should be proud that I kick so much ass in Biology. I will post my gel image on Monday when I can email it to myself.

Lanes: 1--Marker DNA, 2--pRSDNA+XhoI+CIP, 3--gDNA+XhoI. Lanes are denoted in image.