Bio-Lab gel electrophoresis

During gel electrophoresis, four DNA fragments of 800 base pairs (bp), 550 bp, 2200 bp, and 1230 bp are being separated following restriction digest of a plasmid. Put the DNA fragments in order from smallest to greatest distance migrated from the wells.

-2200 bp closest to well -1230 bp close to well -800 bp far from well -500 bp farthest from well

DNA gel electrophoresis equipment

-DNA is placed in an electrical field with one side of the field having a negative charge and the other side having a positive charge. -DNA is negatively charged, so the DNA molecules will move toward the positive side of the field. -Smaller DNA fragments will move farther through the pores in the agarose gel matrix. -The gel box is a container that holds the agarose gel. It includes two electrodes that conduct the current into the buffer chamber from the power supply. Often the cathode (positive) and anode (negative) are color coded red (positive) and black (negative) or indicated by symbols (+ or -). -The power supply generates the current that allows the biomolecules to migrate through the gel matrix. The power supply can be separate from the gel box or may be integrated to create a single unit (as shown in the example below). -The buffer chamber is the part of the gel box that contains the gel submerged in an electrophoresis buffer. This salt-containing solution is what allows the electrical charge to be conducted from the power supply to the molecules in the gel. -The gel sits within the gel tray, which was used to cast the gel from melted agarose. During casting, a comb is inserted into the hot agarose so that as the agarose hardens, empty spaces are left in the gel. These empty spaces will form the wells into which the samples are loaded during electrophoresis. -Because DNA is not visible with the naked eye, a light-reactive stain is often added to the agarose during casting, which allows the DNA to be visualized after electrophoresis. A visualization system like the one shown below will illuminate the gel with UV or blue light, causing the DNA molecules to become visible as bands in the gel. -it is important to use proper protective eye equipment or an orange filter (as shown below) when using a visualization system to prevent exposure to harmful radiation.

PCR is used to amplify a segment of DNA from a crop plant

-after PCR is performed, the PCR product can be visualized using a technique called gel electrophoresis which separates DNA molecules by size -DNA molecules appear as a band on the gel

GMO presence in a crop plant

-if present, PCR product will be produced and appear as a band on the gel -if absent, PCR will not be produced

PCR

-lab technique used to generate billions of copies of a specific sequence of DNA -having many copies of particular sequences allows the ability to read, compare, and manipulate them -can be used to detect the presence of genetic material from pathogens -one application of PCR is to detect the presence of these additional genes (GMOs) in crops -can reveal whether a sample came from a particular species by amplifying a region of the DNA unique to that species (regulators can detect illegal trade if wildlife products)

Gel electrophoresis works by which of the following principles?

-sorting molecules by size -sorting molecules by charge

Gel electrophoresis

-technique that helps scientists sort and visualize small biological molecules based on two properties: size and charge. -it uses a polymer matrix, called a gel, and an electrical current. -in order to see the results of the sorting process, different staining or visualization methods are used to reveal the separated biomolecules, which appear as bands on the gel. -Samples are first pipetted into indentations in the gel, called wells. -negatively charged biomolecules such as DNA are pipetted into wells near the negative (black) electrode. As the current is conducted through electrophoresis buffer and to the gel, the samples move toward the positive (red) electrode -if the charges of the molecules to be sorted are unknown, the wells can be placed in the center of the gel so that the molecules can migrate easily through the gel towards either pole. -to separate DNA or RNA molecules, a polysaccharide (complex sugar) found in seaweed called agarose is typically used. It forms a porous matrix (like a sponge) with lots of holes. The concentration of agarose used will determine how tightly woven the matrix is, and as a result, the size of the pores the molecules travel through. -Since the molecules in the well have different sizes and shapes, they travel through the matrix at different speeds and end up at different distances from the well. All the molecules of a similar size and shape will travel the same speed and distance. -Most biomolecules do not have a visible color, so after the electrophoresis process is completed, a visualization method must be used to observe their final positions -a loading dye is used to follow the progression of electrophoresis, but this dye travels independently of the biomolecules and does not indicate their positions. -fluorescent dyes that bind directly to the biomolecules and fluoresce in the presence of certain wavelengths of light are often used to visualize their positions. -the fluorescent dyes can be embedded in the gel before electrophoresis, or they can be added after electrophoresis is completed.

In gel electrophoresis, DNA is placed in an electrical field with one side of the field having a negative charge and the other side having a positive charge. How will the DNA move in the field?

DNA will be attracted to positive charge and move toward the positive side of the field.

The charge of DNA is _____ due to the phosphate group

negative

Gel electrophoresis can be used in the process of genetic engineering to

verify the size of a piece of DNA so it can be inserted into cells