Sourthern Bloting

Sourthern Bloting

HYPOTHESIS:
The mutation found on the 6th amino acid can lead to the polymerization of the sickle cell hemoglobin resulting in less oxygen concentration in the capillaries.
ALTERED HYPOTHESIS:

DISCUSSION:
The purpose of this experiment was to cut DNA with restriction enzymes and find the size of the DNA fragments. It was also to compare the structure of DNAs from a normal patient with sickle cell anemia. In order to get this done the normal persons DNA was paired with a sick persons DNA with the same restriction enzyme in both. After which a double digest was done with the both enzymes used in both DNA types. The enzymes used were BamHI and EcoRI because I wanted to accommodate the assertion of DNA fragments with complementary DNA. The restriction fragments were then separated using agarose gel electrophoresis. A restriction fragment from the previous lab (Restriction mapping) was used as a probe in order to get the specific DNA species among the DNA on the southern blot. Before the probe was used it was denatured or separated in order to be hybridized effectively to the DNA on the southern blot.The probe was the labeled radioactively. The DNA was then exposed to Ultraviolet light in order for permanent attachment of the DNA to the nylon filter. After which the probe was prehybridized to prepare the probe for hybridization, hybridized which allows the DNA to find and base pair with its complementary strand and filter washed in order to reduce background. The filter was the placed in the Xray cassette for the fragment bands to be seen. A graph was plotted to show this information. On the graph the fragment size was estimated by comparing the position on the gel of the DNA fragment with the position of the gel marker DNA bands. A ruler was used to measure the distance from the starting well to each of the gel marker fragments. The Y axis was labeled the Logarithm of fragment length and the x axis was labeled The distance migrated. A straight line was then...

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