User:Susan Schultz/Notebook/Experimental Biological Chemistry/2011/11/01

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Objective Repeat AuNP synthesis to investigate black fiber formation Use PCR to mutate GFP Description AuNP Synthesis Running 4 AuNP synthesis type reactions in Acetate Buffer, pH 3.6. Reaction 1- 1mL 1.5μM BSA+ 1mL 2.9mM Au remvoing from heat for 10 minutes every 30 minutes Reaction 2- 1 mL 1.5μM BSA+ 1mL 2.9mM Au heating continuously Reaction 3- 1 mL 1.5μM BSA+ 1mL 3MHCl removing from heat for 10 minutes every 30 minutes Reaction 4- 1 mL 1.5μM BSA+ 1mL 3M HCl heating continuously GFP Mutation Mutate GFP to contain cysteine after enterokinase cleavage site. Methods from QuikChange® Site-Directed Mutagenesis Kit Primers:forward (5'-TACgacgatgacgataagtgtcgatggggatccgaattc-3'), reverse(5'-GAATTCGGATCCCCATCGACACTTATCGTCATCGTCGTA-3') In PCR tube combine 5μL 10x reaction buffer, 1μL wild-type GFP DNA, 1μL forward primer, 1μL reverse primer, 1μL dNTP mix, 40μL sterile water, and 1 μL of PfuTurbo Add 50μL Bio Rad Chill Out™ liquid wax Place in thermocycler for 20 cycles of: 30 sec at 95°C, 30 sec at 60°C, 7 min at 72°C Hold at 72°C for ten minutes and leave overnight at 4°C Data Both continuous and interupted synthesis formed black balled fiber. It is possible that they became slightly more purple as the rection progressed. No change in reactions without gold. Notes This area is for any observations or conclusions that you would like to note. Use categories like tags. Change the "Course" category to the one corresponding to your course. The "Miscellaneous" tag can be used for particular experiments, as instructed by your professor. Please be sure to change or delete this tag as required so that the categories remain well organized.