6) Further analysis of the 2 types of seaweeds can be investigated via the procedure of gel electrophoresis. In this case, DNA must be extracted from the seaweed by use of a centrifuge. Upon completion of DNA extraction, the process of protein analysis may proceed. In this particular experiment, the Kjeldahl method was used. The exposed seaweed and non-exposed seaweed are to be digested for approximately 45 minutes in 20 ml of concentrated sulfuric acid in the presence of this catalyst called Kjeldahl. Upon cooling, the digested samples are to be distilled for six minutes, after addition of 50 ml of water and 150 ml of a 0.25% sodium hydroxide solution. The released nitrogen from each sample during distillation is to be collected, the titrated using sulfuric acid. The protein content can then be calculated.
7) Begin the protein analysis process by using gel electrophoresis. This is the separation of nucleic acids or proteins, based on their size, solubility and electrical charge, by measurement of their rate of movement through an electrical field in a gel. The first step is creating a gel (using agarose) that will serve as a medium in which the proteins will separate and move.
8) The gel electrophoresis mechanism consists of 2 glass plates that support the gel, which is bathed in an aqueous solution. Electrodes are attached to both ends, and voltage is applied. Insert each protein solution into the same end of the gel mechanism, ensuring that the lanes have sufficient space between them. Make sure the two solutions do not mix.
9) Observe the movement of the macromolecules through the pores of the gel towards the electrode of opposite charge. This rate of migration through the electric field depends on the molecule’s charge and shape.
10) Upon completion, the separated molecules in each lane will be visible as a series of bands. Study and compare the size and spread between the bands in each lane of the two types of seaweed. The larger molecules will move more slowly due to friction, and will therefore be located near the place of DNA introduction.
RESULTS:
In both regions, the majority of the seaweed was found detached at the root. However, the exposed region proved to display a greater difference between the two categories of seaweed, (either removed at the root or not) than did the non-exposed region. These results may be supported by the idea that the seaweed that was exposed to harsh sea conditions (exposed region) have, over time, adapted to this environment. These seaweed have thus developed stronger hold fasts and roots that enable them to hold tight to rocks, shells and the sea floor at their roots, making it difficult for the seaweed to be broken off at any other region than the root itself. In the non-exposed region, there was not as great a ratio between numbers of seaweed detached at the root or not, suggesting that these roots and hold fasts are not as strong as those found in the exposed region. This is likely due to the fact that the non-exposed region does not subject the seaweed found here to severe, unrelenting conditions as well as the pounding action of the waves.
Force: 1 pound = 4.448 N
EXPOSED REGION (Lions Crescent)
NON-EXPOSED REGION (GREENSLADES ROAD)
Sources of Error:
(1) Since there is more than one species of seaweed growing at Topsail Beach, different types may have different characteristics and therefore may have altered the results if we had not been aware.
(2) The point at which the seaweed was growing within a larger region (exposed or non-exposed) may have had an effect on our data. Those strands that attach to very exposed rocks, in the middle of tidal action would be stronger and thus should not be compared to seaweed that grew in more remote areas, like behind larger rocks.
(3) There are several possible examples of systematic error that may have occurred with the fish scale…
(a) It is possible to have read the scale inaccurately a few times throughout the duration of our experimentation since we performed numerous trials.
(b) The direction in which we pulled on the seaweed with the fish scale may have had an effect on the reading.
(4) The weight or amount of flowers on the seaweed is another issue to bear in mind. A strand of seaweed that is very full of leaves and branches is very different from a strand of seaweed that is just composed of a narrow stalk. Therefore, this may be the cause of some obscure fish scale readings.
(5) The amount of algae present in the water and the direct environment surrounding our specimens may have an effect on breaking down the attachments of the seaweed.
(6) Some seaweed has weak areas, like scar tissues where they had once been broken, and thus they broke at these regions before they did the root.
Metu conducted results - Ocean Sciences Center: MUN
Exposed kelp: Total protein content = 5.3549% (+-0.0419%)
Non-exposed kelp: Total protein content = 5.1934% (+-0.0378%)
Total protein contents of kelp were determined following the Kjeldahl method. Determinatios were carried out in triplicate. 815 mg (+-21 mg) of exposed kelp and 835.5 mg (+-14.5 mg) of non-exposed kelp were digested for 45 minutes in 20 mL concentrated sulfuric acid (H2SO4) in the presence of a catalyst (2 tablets of Kjeltab). After cooling, the digested samples were distilled for six minutes, after addition of 50 mL H20 and 150 mL 0.25% sodium hydroxide (NaOH). The released nitrogen from each sample during distillation was collected in a 50 mL 4% boric acid containing 14 drops of N-point indicator. The protein content was then calculated as follows:
% Nitrogen = (Volume of H2SO4 used to titrate sample x N H2SO4 x 14.0067 x 100) / Weight of sample (mg)
Protein Content = % Nitrogen x 6.25
*Note: 6.25 is a factor (constant) derived from an assumption that 100 grams of protein contains 16 grams of nitrogen (100/16 = 6.25)*
CONCLUSION:
Our hypothesis was indeed confirmed by our experiment. It is apparent that seaweed found in the exposed region had an overall greater strength in terms of ability to cling to the respective attachment point than the seaweed found in the non-exposed region. This confirmation of results can be explained by Darwin’s theory of evolution, that is, organisms can adapt to their environments in such a manner that only the fittest survive. The seaweed found in the exposed region have obviously adapted to their surroundings, developing a more strength-oriented lifestyle. The seaweed typical of non-exposed regions have adapted as well, becoming a species of co-dependant nature. As well as physically testing the seaweed found in these 2 regions, we were able to biologically survey the seaweed by the process of protein analysis. The results indeed proved to be biological evidence that support our actual data from the tests performed. These results showed there was evidently a significant difference between the protein make-up -- the exposed seaweed were composed of much more total protein content, enabling them to successfully thrive in their respective environment.
