ipl-logo

Gel Electrophoresis Lab Report

1709 Words7 Pages
bulb-icon

Recommended: Gel electrophoresis

In biochemistry it is very important to realize that the cells that make us who we are, are all made of specific proteins. These proteins need to be studied and analyzed extremely carefully to understand how certain proteins work in human physiology and in other organisms. In most cases it is important to separate one specific protein so you can better understand that protein’s function. One of the most common ways of separating these proteins is the use of gel electrophoresis. Gel electrophoresis separates proteins based on how big they are. The purpose of this lab was to isolate the protein in cultured E. coli that emits a green glow. The green glowing protein observed is known as Green Fluorescent Protein, or GFP for short. To separate the proteins inside the E. coli, gel electrophoresis and hydrophobic-interaction chromatography was used. Hydrophobic-interaction chromatography is used to separate proteins that are hydrophobic, GFP is hydrophobic therefore did interact with the hydrophobic-interaction chromatography. Knowing that GFP is hydrophobic, it is inferred that it is in the column. To know for sure that GFP was in the elution it was tested under UV light. If it glows green, then it means that GFP was located inside the sample. Once it was ensured that the GFP was in the sample, the electrophoresis …show more content…

Since the proteins should be separated on size only, these components are used. Sodium dodecyl sulfate is used to equalize the charge-to-mass ratio (charge density). SDS works by coating the proteins and disrupting structures to evenly distribute the negatively charged molecules, allowing isolation based on size only. Dithiothreitol breaks cysteine disulfide bonds, disrupting tertiary structures. This allows smooth movement throughout the gel. Last step in preparing the gel includes denaturing the protein at 95 degrees Celsius for 5

Show More
Related
8201 Words | 33 Pages

Figure 20 and 21 casting tray Figure 22 gel box After 1 hr., take out the gel from gel box carefully, place it into the machine, so that the DNA gel electrophoresis can be visualize under UV light. Figure 23 gel documentation system, used to visualize gel electrophoresis with UV light NMR spectroscopy First of all, 3 samples were prepared, peptide in SDS, DPC, and buffer. The sample temperature was maintained at 298 K, prepared by supervisor and H(hydrogen) in SDS and DPC micelle was replaced with D(deuterium) , so that in proton NMR, peptide won’t be interfered by H in micelle. amount of peptide in sds and dpc

Read More
1764 Words | 8 Pages

1.1 Abstract The purpose of quantitative analysis of protein using a spectrophotometer is to measure the concentration of proteins in a given sample. The experiment is conducted by laboratory method (Biuret Test) and using spectrophotometer to analyze the absorbance of reactants at 540 nm, hence determining the concentration of the proteins in a given sample. The purpose of stopped enzyme assay to study B-galactosidase is to determine the effect of temperature and concentrations of substrate on enzyme activity.

Read More
811 Words | 4 Pages

Figure 2a Figure 2b Figure 2a represents the gel fully run through gel electrophoresis for thirty minutes and figure 2b represents the expected results from the gel electrophoresis run. As seen by figure 2b, there are bands present in all lanes except for land three which is the wild type digested DNA sample. It is very faint, but in lane three of figure 2a, there appears to be a slight band apposed to the

Read More
914 Words | 4 Pages

The process of transformation is used often to evolve or change a cell by implanting new genes which can safely alter the genes already located in the organism we are trying to change. In this lab, the goal was to transform E coli. to glow in the dark, or in other words to make transform it and give it the ability to be bioluminescent. Bioluminescence is one of nature’s most useful and stylish abilities. In many species the ability to create a glow based off the DNA located in the cell is used in a variety of ways-

Read More
700 Words | 3 Pages

The secondary antibody (anti-rabbit HRP labelled) used is conjugated with an enzyme Horse radish peroxidase which then binds to its substrate tetramethyl-benzidine (TMB) to produce a blue color indicating the presence of lysozyme. SDS-PAGE gel electrophoresis is a process which separates proteins based on molecular weight. The purpose of this method is to separate out the lysozyme by its weight. The weight is known to be 14.6Da. To view the protein separation, the gel was placed into coomassie blue to develop a series of bands to detect the protein by its

Read More
565 Words | 3 Pages

In this experiment, we had purified LDH isozyme from the clarified homogenate via the ammonium sulfate precipitation, affinity chromatography and size exclusion chromatography. Based on the result shown on the LDH table and on the SDS page, the most effective LDH purification step is the affinity chromatography. Even though the affinity purification step recovers the least amount of LDH (only 3% of LDH from the previous 65% cut), it shows the most purified enzyme without any contaminating proteins on the SDS PAGE. Since the effectiveness of purification is determined by both the recovery rate and the purity rate, other LDH intermediates do meet the qualifying condition. The 65% cut pellet has the highest recovery rate, 84% of the LDH of

Read More
741 Words | 3 Pages

SDS-PAGE can be used for many different uses such as; finding important proteins in a sample and figuring out how much each protein is in a sample (Experimental Biosciences). Coomassie staining is used on SDS-PAGE gels after proteins get separated by SDS-PAGE machine, in which the SDS-PAGE gels get immersed in coomassie stain and the extra amount gets rinsed off with solvent. When the staining of SDS PAGE is completed is should show blue bands on the clear background of the SDS PAGE gel (BIO RAD). Western blotting is a lab procedure that is used to find specific proteins in a mixture

Read More
502 Words | 3 Pages

Agarose gel electrophoresis is an easy and common technique of separating and analyzing DNA. The main objective of this lab is to find the sire of the offspring using gel electrophoresis. Gel electrophoresis is used in laboratories to isolate charged molecules like DNA, RNA, and particular proteins according to their specific size. The charged molecules travel through the gel when an electric current is spread across it. The electric current is applied across the gel so that the ends of the gel have a positive charge and the other end has a negative charge.

Read More
153 Words | 1 Pages

The molecular-based method used by the scientists to differentiate between the species of daisies was gel electrophoresis. Gel electrophoresis is a technique used to separate charged molecules such as DNA, RNA, and proteins by their sizes. To begin with, samples are loaded at the top of the gel in the same horizontal position. Once the samples are loaded, an electric current is applied, which allows the charged molecules to migrate to the opposite end. For example, negative charged molecules will migrate towards the positive end.

Read More
535 Words | 3 Pages

Smaller molecules can diffuse into the beads and move more slowly down the column. The process of extracting one material from another by washing with a solvent is called elution; they are three ways isocratic elution, gradient elution, and stepwise elution. In this experiment the process of isocratic elution will be uses, where the same mobile phase is used throughout the elution and allows separation of molecules that differ significantly in size. By varying the degree of cross-linking the gels are optimized for different molecular weight ranges. The purpose of this experiment is to analyze and identify specific proteins from a mixture of proteins using the purify technique of size exclusion chromatography.

Read More
1018 Words | 5 Pages

Their visualisation was performed using Coomassie brilliant blue (G-250). GS-800TM Calibrated Densitometer (BioRad, California, USA) was used to scan the gels and the resulting image file was assayed using the Quantity One analysing software (BioRad, California, USA). Molecular masses were estimated according to Precision Plus Protein unstained standards (BioRad, California, USA) which contains ten highly purified recombinant proteins with molecular masses from 10 to 250

Read More
1284 Words | 6 Pages

When AC is used to purify and separate large biomolecule from a complex mixture, those techniques aspects have to be consider. The main function of matrix is allowed ligands to bind it tightly; regardless which type of matrix was used in the experiment, several perspectives have to be considered when choosing a proper support matrix, such as chemical stability and chemical inertness. There are numerous techniques to separate protein based on its natural property, such as, separate protein based on various mass by ultracentrifugation or based on charge and mass ratio by SDS-polyacrylamide Gel electrophoresis (SDS-PAGE). Those purification methods would disclose some information about target protein; however, the result is less accurate and little vague to cause false positive. Compare to other purification method, the AC had various unique advantages, for instance, it recognized and purified biomolecule based on its biological function or chemical structure, and purification process took less time-consuming and accomplished with high recovery rate of target

Read More
813 Words | 4 Pages

The Cu1+ then react to bicinchoninic acid assay forming a purple water soluble complex. Moreover, the total volume of protein concentration can be measured by the colorimetric technique which will change the colour of sample solution from green to purple in proportion to protein concentration. SDS-PAGE electrophoresis Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE electrophoresis) is probably the most common analytical technique widely used to separate biological molecules, usually a nucleic acid or protein based on their electrophoretic mobility. The motility is a function of conformation, the length of their peptide chain and charge of the molecule. Depending on their size, small biomolecules move faster and more easily fit through the pores in the gel than larger ones.

Read More
719 Words | 3 Pages

CZE, also known as free solution capillary electrophoresis, is a separation technique that predominantly takes into account the ratio of the particle’s charge to mass, where those with large charge to mass ratio separate from the rest first; therefore, the larger the ratio, the quicker the separation. In addition to the electrophoretic mobility of the molecules, CZE is heavily dependent on the application of constant field strength throughout the capillary and on the pH of the buffer solution. CZE is an excellent choice of technique to employ in cases where there are very small pI (isoelectric point) differences in protein

Read More
1251 Words | 6 Pages

Finally, the amplified DNA regions are compare using a gel. DNA Profiling

Read More

More about Gel Electrophoresis Lab Report

Open Document