Nucleic Acid Chromatogram O O Nucleic acid chromatogram with nucleic acids circled in pencil 00
Please explain the results and conclusion based on the results of these two chromatograms (one for proteins and one for
For the proteins chromatogram, the Xs on the line from left to right are:
- unhydrolyzed protein
- hydrolyzed protein
- alanine solution
- histidine solution
- aspartic acid solution
- lysine solution
- methionine solution
- a solution containing an unknown amino acid (it will be 1 of the 5 amino acids found in microfuge numbers #3 to #7 above)
For the nucleic acid chromatogram, the Xs on the line from left to right are:
- unhydrolyzed nucleic acid
- hydrolyzed nucleic acid
- adenine solution
- cytosine solution
- uracil solution
- adenine-cytosine-uracil solution
#1 and #2 for both proteins and nucleic acid chromatogram was prepared with yeast cells and centrifugation.
- Yeast was grinded down with sand and then TCA (trichloroacetic acid)
The yeast contains yeast cells held together with a starchy binding material. Glucan (a cellulose-like polysaccharide) is present in the yeast cell walls, and glycogen, proteins and nucleic acids are present in the cytoplasm. Grinding the cells with sand will rupture the cell walls and cell membranes. TCA (trichloroacetic acid) was then added to the mixture. Polysaccharides such as starch and glycogen are soluble in TCA, and they will go into solution in the TCA. Proteins and nucleic acids are insoluble in TCA, and they will remain in suspension.
- Centrifuge, discard supernatants and keep pellets
- Add NaCl to pellets
- Now have: 4 tubes with each containing proteins and nucleic acids
- Boil 4 tubes, centrifuge, decant into beaker
- Now have: 1 beaker with nucleic acid and 4 tubes with proteins
- Add ethanol to beaker with nucleic acids
- Centrifuge suspension and add sulfuric acid to new pellets
- Transfer liquid into 2 glass tubes
- Boil 1 glass tube and leave other 2
- Boiled tube is hydrolyzed nucleic acid and unboiled tube is unhydrolyzed nucleic acid
- Neutralize content of both tubes with Ba(OH)2
- Refrigerate content and chromatography is then done to separete and identify proteins and nucleic acids
Rf calculation = distance from origin (line with the Xs) traveled by substance (where the substance ends) / distance from origin traveled by solvent (the line at the top right)
All calculations are done in cm and it’s fine that it’s measured from the screen (no need to print). There’s also no need to determine all the Rf values but discuss the results and conclusion based on the results. For both protein and nucleic acid chromatogram, are there any results that are not expected? If so, what should this result have actually looked like? Why might this have happened?
High Rf value means that solute was attracted to the solvent (which was moving)
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