ANALYSIS
The isolation of RNA from yeast involves heating with NaOH which served to disrupt the cell membrane and lyse the cell extracting the nucleic acids. The NaOH also increases the pH level of the solution resulting in the denaturation of contaminant proteins, inactivates nucleases which can degrade RNA. Heating helped loosen the cell membrane by increasing the inetic energy of the lipid molecules, maing it release more RNA. The mixture was filtered and centrifuged to get rid of the denatured proteins, lysed lipid membranes and other contaminants. The purpose of the addition of glacial acetic acid was to lower the pH level to help denature more proteins, prevented alali RNA hydrolysis, ensuring that the desired RNA was not degraded. The mixture was decanted and centrifuged repeatedly to eliminate the precipitated proteins. The reason why the supernate was suggested to be !"m# or below was to increase the RNA in the solution and for it to be easily isolated later on. $thanol was added to lower the dielectric constant of the solution and reduced the solubility of RNA causing it to precipitate from solution. H%l was added to protonate the phosphate groups in nucleic acid bacbones, minimi&ing the charge repulsions between molecu molecules les and helped helped aggreg aggregate ate and precip precipita itate. te. %entrif %entrifugat ugation ion also also separa separated ted the RNA precipit precipitate ate from from the unneeded supernatant. 'oth washings from ethanol and ether removed any lipid residues and other non(polar contaminants.
Acid Hydrolysis is a chemical process in which acid is used to convert cellulose or starch to sugar. )n this experiment, experiment, the RNA yields yields products products of hydrolysis hydrolysis which is the Nucleobases Nucleobases *A,+, *A,+, - %, /ugar *0(ribose *0(ribose,, and 1hosphate.
The hydrolysate increases amino acid oxidation, it is beneficial for protein synthesis as it reacts 2uicly to the reagents and chemical used maing the reactions faster than the unhydroly&ed RNA. This was evident in our observations throughout the experiment when we tested each using the two different RNA. The products in the 'enedict3s test both tested positive *4 when they reacted with the hydrolysate, and the unhydroly&ed in separate test tubes. Orcinol Test or also nown as the test for 1entoses only produced one positive *4 product when tested with the unhydroly&ed5 1urine bases yielded an acidic solution in the hydrolysate, while it yielded a basic solution in the unhydrol unhydroly&e y&ed. d. The last last experim experiment ent conduct conducted ed was the )norg )norgani anicc 1hosph 1hosphate ate test, test, which which produc produced ed positi positive ve *4 solutions in both the hydrolysate and unhydroly&ed.
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'ene 'enedi dict ct33s Test The 'enedict3s test, in order to be classified as having positive *4 solutions, should yield colors of green or red indicating the presence of reducing sugars5 The colors that the solutions produced in our experiment were light green *hydrolysate and unhydroly&ed indicating the presence of a very low reducing reduc ing sugar.
6
Test for 1entoses *Orcinol Test /olutions having the presence of pentoses with orcinol reagent *7,8 dihydroxytoluene will always yield a blue(green solution. Only one in the experiment produced a positive *4 solution where the unhydroly&ed was involved5 the other solution with the hydrolysate produced a yellow(brown *almost dar complex, which was due to prolonged heating of hexoses that yield hydroxymethylfurfurals, which reacts with the orcinol giving the product a yellow(brown color.
7
1urine 'ases 9ree purine bases may be separated from the protein nucleosides by precipitation with the help of silver nitrate *AgNO7. The hydrolysate tested acidic with precipitate *formed 2uicly which is a purine complex with Ag4 ion forms, and the unhydroly&ed that tested as basic with precipitate that was viewed und er the microscope.
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Test for )norganic 1hosphate There is presence of inorganic phosphate. On warming with the ammonium molybdate in presence of HNO7, inorganic phosphate is precipitated as yellow ammonium molybdate. 'oth solutions tested as positive *4 yielding a yellow color.
The difference between the hydrolysate and the unhydroly&ed RNA was how the reaction taes time5 Reactions that too place with the presence of the hydrolysate were faster than the experiments under unhydroly&ed, this was due to the fact that the hydrolysate went under acid hydrolysis.
Terms: 0ecant( gradually pour *li2uid, typically wine or a solution from one container into another, especially without disturbing the sediment. %entrifuge( is a piece of e2uipment that puts an ob;ect in rotation around a fixed axis *spins it in a circle, applying a potentially strong force perpendicular to the axis of spin *outward. /upernatant( denoting the li2uid lying above a solid residue after crystalli&ation, precipitation, centrifugation, or other process.
CONCLUSION
Ribonucleic acid *RNA functions in converting genetic information from genes into the amino acid se2uences of proteins. )n this experiment, RNA was isolated from yeast *Saccharomyces cerevisiae) by heating the active dry yeast with alaline NaOH. This method of RNA extraction involved the disruption of the cell membrane and subcellular nucleus to brea open and discharge the nucleic acids. RNA was extracted from associated proteins with H%l extraction and was treated with ethanol and ether to remove lipids.
REFERENCES • • • • • •
http://www.pocdscientifc.com.au http://www.slideshare.net/mobile/kevbalda/report-exp-6-and--dna-and-rna http://publications.lib.chalmers.se/records/fulltext/!666"#.pdf http://www.nou.edu.n$/uploads/%&'%(&*/pdf/pdf#/+,&##!.pdf http://umanitoba.ca/+iolo$y/+,&*!#/lab#/biolab#(#.html http://www.0bc.or$/content/!1"/!/#!!.full.pdf
