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==Description== | ==Description== | ||
Two separate procedures were done. | |||
1)A culture media of E.Coli was prepared for the protein expression. | |||
-0.875 g of Lysogeny Broth and 35 ml of water were added to four separate 250mL flasks and by using a piece of aluminum foil top of the each flask was covered. | |||
-Flasks were placed to autoclave machine for 50 minutes. The temperature was 121 F. | |||
-After being autoclaved for 50 min, they were let to be cooled. | |||
-0.450g of ampicilin was mixed with 4.5 ml of water. | |||
-35uL of the ampicilin mixture was added to each flask. | |||
-By using sterile methods, bacteria was transferred to each flask with ampicilin mixture. | |||
-The flasks were placed in the uncubater overnight. The incubater was at 37°C. | |||
2)GFP containing a cysteine was prepared by using PCR. | |||
-Online webpage,( http://www.basic.northwestern.edu/biotools/oligocalc.html) was used to have a better understanding of PCR and find the forward and reverse primers necessary for the experiment. | |||
Forward: 5' GAC GAT GAC GAT AAG GAT CGA TGG GGA TCC GAA 3' | |||
Reverse: 5' TTC GGA TCC CCA CCA TCG ATC CTT ATC GTC ATC GTC 3' | |||
-A solution of 5 uL of 10X Pfu reaction buffer,1 uL of PfuTurbo DNA polymerase,1 uL of dNTP mix,1 uL of template DNA, 1.25 uL of primer 1 and 1.25 uL of primer 2 was prepared. | |||
-Before placing the solution in the thermocycler at 95°C for 30 seconds, 25 uL of BioRad Chill out wax, dark pink, was added to the top of the solution in order to prevent the evaporation of the solution. | |||
-The solution with the wax was kept in the thermocycler, which was at 4°C, for 24 hours. | |||
==Data== | ==Data== | ||
Revision as of 11:35, 26 September 2011
 Biomaterials Design Lab
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Entry titleObjectiveDescriptionTwo separate procedures were done. 1)A culture media of E.Coli was prepared for the protein expression. -0.875 g of Lysogeny Broth and 35 ml of water were added to four separate 250mL flasks and by using a piece of aluminum foil top of the each flask was covered. -Flasks were placed to autoclave machine for 50 minutes. The temperature was 121 F. -After being autoclaved for 50 min, they were let to be cooled. -0.450g of ampicilin was mixed with 4.5 ml of water. -35uL of the ampicilin mixture was added to each flask. -By using sterile methods, bacteria was transferred to each flask with ampicilin mixture. -The flasks were placed in the uncubater overnight. The incubater was at 37°C.
-Online webpage,( http://www.basic.northwestern.edu/biotools/oligocalc.html) was used to have a better understanding of PCR and find the forward and reverse primers necessary for the experiment.
-A solution of 5 uL of 10X Pfu reaction buffer,1 uL of PfuTurbo DNA polymerase,1 uL of dNTP mix,1 uL of template DNA, 1.25 uL of primer 1 and 1.25 uL of primer 2 was prepared. -Before placing the solution in the thermocycler at 95°C for 30 seconds, 25 uL of BioRad Chill out wax, dark pink, was added to the top of the solution in order to prevent the evaporation of the solution. -The solution with the wax was kept in the thermocycler, which was at 4°C, for 24 hours. Data
NotesThis area is for any observations or conclusions that you would like to note.
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