User:Samiye Yaman/Notebook/Lab 571/2011/09/20

From OpenWetWare

< User:Samiye Yaman | Notebook | Lab 571 | 2011 | 09
Revision as of 13:35, 26 September 2011 by Samiye Yaman (Talk | contribs)
(diff) ←Older revision | Current revision (diff) | Newer revision→ (diff)
Jump to: navigation, search
Biomaterials Design Lab Main project page
Previous entry      Next entry

Entry title

Objective

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

  • Add data and results here...

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.



Personal tools