The gel electrophoresis of the protein extract, extraction buffer, and ADH was done. The second part of the experiment went smoothly without any mistakes. The proper guidelines provided by laboratory personnel and in Brand (7) was followed. Protein bands observed in Lane #5 were not perfectly horizontal, instead, protein bands were in “wavy” like configuration. This made it difficult to measure the distance of each bands from the origin (Fig. 4). Several protein bands were noticed in the lanes containing protein extract. Lane #3 did not produced any protein bands. The extraction buffer did not contain any proteins; extraction buffer served as the controlled experiment. Since, extraction buffer was mixed with protein extract.
The gel electrophoresis done was compared to other research groups. In comparison, other group’s gel was not completely destained. Their gel for that reason did not produce distinct and attractive patterns of protein bands. To produce distinct contrast of protein bands, the gel was destained many more times using the tissue paper and destaining solution. More time the destaining solution was used, protein bands became more distinct against the gel background.
The protein electrotransfer was prepared on the other half of the slab gel unit. The MilliBlot electrotransfer unit was used to transfer the protein bands from the gel to the PVDF membrane. Protein would precipitate from the gel to the PVDF membrane based on electrical charge distribution. Cathode buffer was used to soak both the electrotransfer membrane and the corresponding gel. Cathode buffer used was not measured. Proper procedures were followed during this part of the experiment. The electrotransfer membrane and gel were not correctly placed in MilliBlot unit. This caused ineffective contrast of protein bands on the membrane. The protein bands on the membrane were not clearly distinct; they faded into the membrane background. Other groups’ membrane produced bands that were clearly noticeable.
Protein bands were compared among lanes. Looking at Fig. 3, 4 and 5, protein bands distance from their origin can be compared. ADH protein bands were found closely related to the bands in Lane #2 and #4 (protein extract). This can suggest that some of the protein found in horse and cow have similarity based on their molecular weight. For this reason, one can use the Fig. 3a in able to approximate the molecular weight of the proteins separated from the protein extract. Just because the bands weigh the same does not make them similar in molecular and structure composition. Other groups’ Rf values for lane #1 and #5 were similar. It was difficult to figure exactly where on the protein band to measure the distance from the origin. Since each protein bands were only few mm long, the measurement of each protein band was take from the center of that band to the origin.
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