Monday, August 6
Colony PCR of U construct using four new colonies and the same positive Z control has once again resulted in no successful colonies. However, the PCR has reaffirmed that the control works, which is a good sign.
We also did a PCR of the color constructs in order to form our BioPaint set. [PICTURE BELOW]
Since our T plasmid and Z construct ligations have not been going well, we started over by preparing new cultures of Z and T, running them on a gel, and then gel purifying the results. [SEE PICTURE]
We began 5 ml cultures for the good Z construct with TOPO and for one of the U ligation colonies. We will use the Z culture for a glycerol stock, and digest the U culture to see if our ligations actually succeded.
Tuesday, August 7
Today, we digested the U ligation construct by cutting it, in two tubes, with AgeI-HI and NcoI. A double cutter, the AgeI-HI would excise out the insert, whereas NcoI cuts once. The results, shown below, indicate [PICTURE + FINISH]
We also PCR'd up the Z construct, U construct, and Z+TOPO constructs using KlenTaq.
Wednesday, August 8
This morning, we ran the three PCRs from yesterday on a gel. [RESULTS OF GEL]
We took the working Z and U constructs, digested them with EcoRI (E) and PstI (P), and ran them on a gel. [PICTURE]
Finally, we started cultures of our Z ligation, control, and a zeaxanthin culture to use tomorrow to test whether our genes are working properly.
Thursday, August 9
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