BS111LFinalReview - Dan McAree BS111L Written Final Review...

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Dan McAree BS111L Written Final Review Scientific Method Hypothesis- educated guess Manipulated variable- aka independent or treatment variable- the parameter you vary or manipulate between treatments Response variable- aka dependent variable- the parameter affected by your manipulations Controlled variables- all parameters that you keep constant between treatments A control is a standard against which experimental results can be compared. Positive controls confirm that your assay is working properly and show you what a positive result actually looks like. Negative controls confirm that you are not getting false positive results and show you what a negative result looks like. Biological Molecules Module Benedict’s Test for reducing sugars (carbohydrates): Pos= red precipitate/glucose. Neg= blue /distilled water Biuret’s test for protein detection (qualitative): Pos= violet /albumin. Neg= blue /distilled water Bradford’s test for protein concentration (quantitative): Pos= intense blue / albumin. Neg= not blue (i.e. orange)/ distilled water SDS-PAGE for protein fingerprinting (proteins): Pos/neg: (proteins work, non-proteins don’t) Sudan III Test for fats (lipids): Pos= orange /vegetable oil. Neg= pink / distilled water Iodine Test for Starch (carbohydrate): Pos= black /starch. Neg= brown / distilled water Glucose + fructose (monosaccharides) -- sucrose (disaccharide) Proteins= polypeptides What SDS does to the proteins: Extracting the proteins from tissue, we load our extracts onto a gel made of polyacrylamide and expose the gel to an electrical field. SDS (sodium dodecyl sulfate), a detergent, is added to the gel to help give the proteins a uniform negative charge. What heating does to the proteins: Incubating the proteins at 95degress Celsius denatures the proteins which are in the liquid. Why the proteins move through the gel: Charged proteins, when placed in an electrical current, will move toward either the anode or cathode depending on their overall charge. So placing a sample of proteins in a semi-solid gel and exposing them to an electrical current will make the proteins move through the gel. Relationship between protein size and migration through the SDS-PAGE gel: As the different molecules pass through the pores of the gel, smaller molecules will travel through the pores more easily, quickly, than larger molecules- separating the molecules in the gel by size. The resulting profile of protein bands in the gel is called a protein fingerprint. When comparing an unknown protein band to kaleidoscope what does that tell you about the unknown: Kaleidoscope is a sample of proteins of known molecular weights so comparing allows you to determine the actual molecular weight of an unknown protein.
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Electrophoresis (SDS and agarose) – in agarose gel electrophoresis, the DNA sugar- phosphate backbone is naturally negatively charged so it moves through the gel toward the positive end, the cathode. In SDS-PAGE, you have to heat the proteins to denature
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BS111LFinalReview - Dan McAree BS111L Written Final Review...

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