Dna gel electrophorosis
DNA, Deoxyribonucleic acid, is a double stranded, helical nucleic acid molecule which determines inherited structure of a protein. The "steps" are made of bases: adenine, guanine, cytosine, and thymine. The sides are sugar and phosphate molecules. Restriction enzymes are enzymes that cut DNA at restriction sites, leaving fragments blunt or sticky. The restriction fragments are separated using a technique called DNA has a negative charge so when an electrical charge is applied it makes DNA move to the positive side. DNA is placed in agarose gel. Smaller fragments move faster. The purpose of this lab is to separate DNA fragments using gel electrophoresis. Hind III cuts AAGCTT between the two irst A's. EcoRI cuts at GAATTC between the G and the A. Hind III and EcoRI both make sticky ends. Our results for this lab were EcoRI separated into five fragments. Hind III separated into four fragments. The control only had one fragment.
We dipped the pipet trough the surface of the buffer over the wells and expelled the contents. The box was filled with tris-borate-EDTA buffer so it covered the entire surface of the gel. We waited ten minutes for the gel to solidify. The power supply was turned off after the bands migrated near the end of the gel and the top of the electrophoresis chamber was removed. We used Hind III, EcoRI, and a controlled enzyme. The top of the electrophoresis chamber was closed and electrical leads were connected. (See chart A and figure 1-1 for distances) Discussion: The purpose of this lab was to see how gel electrophoresis separates DNA fragments. The amount of lines show how many fragments there is in the DNA. The micro pipet was used to load the lambda EcoRI, lambda Hind III, and lambda only into the wells. Procedures: We sealed the ends of a gel casting tray with masking tape and inserted a comb into the slots. We removed the gel from the gel casting tray and examined it under a light box and compared it to the ideal gel (figure1-2). Loading the DNA was difficult and if you weren't careful you could rupture the wells which ruined the lab.
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