Team:BYU Provo/Notebook/CholeraDetection/Springexp/Period2/Dailylog

5/13/13

Today we ran our digested CRO insert on the low melt gel, and under the UV light we saw the band we were looking for. Clarisse excised the band and we performed a ligation reaction according to the protocol and using the vector that we had already run on a low-melt gel last Thursday. After incubating for 30 minutes at room temperature, we performed a transformation into DH5alpha. The selectable marker of pLAT is ampicillin.

We ran our low melt gel with our Cro insert. After 45 minutes, we put the gel under UV light and cut out the insert. We then performed a ligation of our vector and cro insert. After 30 min, we did a transformation and inserted our newly ligated plasmid into DH5alpha.

KK, KP

5/14/13

(Thursday) When we dropped by today two of our plates (out of four) were contaminated. The two contaminated plates, we noticed, were the two plates that were old. The other two plates had a few colonies, but the was not a significantly greater number of colonies in our plasmid + insert + E.Coli plate than there were in our control plasmid + E.Coli plate. Kelton went ahead and streaked the few colonies we had to singles. We can PCR-verify if any of them have the plasmid. Meanwhile I am going to redo the transformation today. I will plate 4 plates: one 50 microL test, one 100 microL test, one 50 microL control, and one 100 microL control.

KK, KP

5/15/13

On Saturday we checked our bacteriophage spot test for plaques, but we saw none. Our bacteriophage lambda purification procedure failed. Dr. Grose thinks that inducing lambda to lysis by 43 degree heat shock must not have been sufficient stress. Since we know H202 does induce our phage, we're going to attempt to isolate bacteriophage lambda by doing a top agar plaque assay with a drop of hydrogen peroxide, then bathing the top agar in LB, sucking it all up and then purifying the bacteriophage by chloroform lysis procedure.

Clarice has been working on submitting parts to the registry. She has cloned the Ydiv gene into the iGEM backbone to submit the part; we purified the plasmid for submission.

KK, KP, CH