Team:University of Ottawa/29 July 2008

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=Today in the lab=

Chris

 * PCR Amplification of PTP2
 * <li> Began running experimentation alongside Matt to increase chances of success
 * <li> Ran 5 samples, including two water controls as per Cory's request.
 * <li> Master mix: 50 uL Buffer, 5 uL dNTP, 12.5 uL F60 and F61, 2.5 uL DNA (at 25 ng/uL), 142.5 uL water.
 * Digestion of PSSA42
 * <li> Ran five samples and one water control
 * <li> Per tube: 3 uL water, 3 uL Buffer 3, 3 uL BSA, 1.5 uL XhoI and BamHI, 16 uL DNA template.
 * <li> Incubated at 37 C for one hour
 * PCR Cleanup
 * <li> Used PCR cleanup kit to purify PTP2
 * <li> Measured absorbance of resulting DNA samples. The concentrations were found to be very low; PCR did not work. It was later determined that DNA template was not added, by accident.
 * PCR Amplification of PTP2
 * <li> Ran PCR with same constraints as previously. Let run overnight.

Tammy and Dan

 * 0.8% Agarose Gel Electrophoresis of T123 PCR Products
 * <li> Each PCR reaction tube (50 &mu;L) was divided into 2 and ran on 2 separate wells to decrease DNA load and to ensure identification of appropriate band size


 * Agarose Gel Extraction
 * <li> 24 bands were cut from the gel using minimal UV exposure
 * <li> 6 gel bands were pooled into 1 column for a total of 4 aliquots of purified T123 DNA

Matt

 * Inoculation
 * <li> Inoculation of pSSA42 turned out nicely, control was clean - performed a miniprep from inoculation.
 * Transformation
 * <li> Plates of transformation in competent cells did not yield any colonies - control was clean.
 * Digestion
 * <li> Digestion of pSSA42 was performed with BamHI + xhoI.