Notebook
Day
1 [
19/07
]
•
Learned how to safely use pipettes.
•
Extraction of target plasmids.
•
Amplification of fragments by PCR.
•
Agarose gel electrophoresis of fragments.
Day
2 [27/07]
•
Cutting of the agarose gel and the extraction of the gel.
•
Double enzyme digestion
•
Formation of recombinant plasmids
Day 3
•
Preparation of LB solid culture medium from the liquid medium.
•
Transformed
the recombinant plasmids.
Day
4
•
PCR identification of transformants
•
Inoculate positive transformant seed solutions to LB culture medium
plates.
•
Processed the resulting pictures for the experiments (gel, petri dish
with bacteria etc.)
Day
5-6
•
Expression of positive transformant containing plasmid A and C.
•
Prepared competent cell (BL21(DE3) containing plasmid B).
•
Transformed plasmid D to competent cell BL21 containing plasmid B.
•
Prepared SDS-PAGE gel.
Day
7
•
Obtain
cell
lysate
by ultrasonic crushing machine.
•
Used Ni2+ column purification method to purify the cell lysate.
•
Used SDS-PAGE gel electrophoresis to identify protein samples.
•
Stained the gel after electrophoresis for observation.
Day 8
•
Isolated bacteria from E.coli and inoculated them to petri dishes.
•
Destained the SDS-PAGE gel.
•
In vitro cleavage of purified proteins.
•
Proceeded with agar diffusion growth inhibition assay.
Day 9
•
Analyzed the results of the whole experiment
•
Uploaded basic and composite parts to the iGEM website.
Day 10-11
•
Proceeded with agar diffusion growth inhibition assay.
Day 12
•
Prepare samples and sent to SubCat for agar diffusion growth
inhibition assay.