Electrophoresis of Proteins from Cauliflower Fractions by SDS-PAGE Laboratory Summary Electrophoresis is a technique where molecules are separated according to their physical properties such as size‚ charge‚ and/or shape. Charged proteins are commonly separated in this matter using PAGE (polyacrylamide gel electrophoresis) to identify individual proteins present in samples. In this lab‚ SDS-PAGE was used. SDS-PAGE separates charged proteins primarily by size because the ionic detergent sodium dodecyl
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experiment‚ an absolute standard was calculated using BSA‚ so that the concentrations of the other unknown protein solutions can be determined (Lambert et.al‚ 2011).The different assays used for this protein quantification were Lowry‚ Bradford (Coomassie Blue) and UV direct. Protein assays help to determine the amount of desired particle
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Objectives Protein Isolation: Protein isolation for a western blot uses detergents and mechanical force to separate seeded cells from their container. Eukaryotic cells are attached to the surface of a flask by cadherins. In the past‚ we’ve separated the cells from the flask by breaking these bonds with a protease‚ but in order to keep the proteins intact‚ a different method needs to be used to extract the proteins. In protein extraction for a western blot‚ we use detergents to lyse the membrane
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amounts of Bovine Serum Albumin (BSA). Varying amounts of BSA at a concentration of .5µ/µl were mixed with the water and 1mL of coomassie dye. The equation y=0.0259x +0.0511 represents the best fit line for a data set taken from µg of protein equals two to ten µg. Optical density (OD) was surveyed using a Mach V visual spectrophotometer at 595. Worktable for Coomassie Assay Tube # | Sample ID | Water (µl) | Sample (µl) | Protein (µg) | OD @ 595 | 1 | BLANK | 1000 | 0 | 0 | 0 | 2 | STAN
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Calculations are as followed: Sample (Tube) A was prepared with pure dH2O since the concentration of protein was 0 μg/mL. Upon addition of Coomassie Blue G-250 and the completion of the incubation period it was noticeable that solutions of higher concentration had a deeper color blue opposed to those with a lower concentration. The absorbance range for the standard curve (figure 1) was determined
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Background: Lipid and protein composition varies in tissue types. Results: When comparing liver‚ muscle‚ and adipose tissue‚ adipose tissue had the highest percent lipid and liver tissue had the highest percent protein. Conclusion: Based on analysis of the gels‚ the liver appeared to have the most variety in proteins when comparing liver‚ muscle‚ and adipose tissue. Significance: In order to determine the composition of a tissue‚ specific macromolecules can be extracted‚ quantified‚ and analyzed
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increase the absorbance readings‚ and skew the accuracy of the protein concentration. BCA does not react to contaminants such as detergents (2). In the Bradford assay in the readings are usually caused by elevated levels of detergent (3). When Coomassie Brilliant Blue G-250 binds to proteins‚ it causes a shift in the absorption maximum of the dye from 465nm (red) to 595nm (blue) in acidic solutions (2). Things that cause a change in the solution from acidic too basic will skew
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Introduction Transformation is a genetic alteration of a cell resulting from the direct uptake and incorporation of exogenous genetic material from its surrounding through the cell membrane. The arabinose operon changes AraC from a repressor to an activator; in this experiment the pGLO plasmid has been designed with a modified operon so that in the presence of the arabinose the bacterial cells which have been transformed by the pGLO plasmid will fluoresce due to the production of GFP. SDS-PAGE
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Beta-Galactosidase and Western Blot 0 1. EMG 9 and EMG 26 contain strain _lac-_(I- Z+ Y+) and strain _lac -_ (I+ Z- Y-)respectively.Three genes huddled together on the chromosome are required for two strains of _E.coli_ to utilize lactose.Consisting of three genes‚ namely‚ _lacZ_‚ _lacY_ and _lacA_‚ the _lac_ operon orderly handles these genes to code specific enzymes necessary for the metabolism of lactose. The genes _lacZ_‚ _lacY_ and _lacI_ would code for beta-galactosidase‚ galactosidase permease
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In this lab we employed various assays utilizing a biuret reagent‚ coomassie brilliant blue reagent‚ and ultraviolet light in order to determine the identity of six unknown solutions and the concentration of a bovine serum albumin sample. We were given three samples that lacked protein‚ and three samples containing proteins‚ and using a spectrophotometer we assessed the amount of light absorbed versus the light transmitted‚ based on the principles of the Beer-Lambert Law. The three proteins used
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