Lec6 | Gel Electrophoresis Flashcards
Why is gel electrophoresis performed after PCR?
Gel electrophoresis is performed to visualize and confirm that the intended DNA fragments were successfully amplified.
How does gel electrophoresis separate macromolecules like DNA?
Gel electrophoresis separates molecules based on their size and charge as they move through a gel matrix under an electric field.
Why does DNA always migrate toward the positive electrode in gel electrophoresis?
DNA is negatively charged, so it is attracted to the positively charged electrode during gel electrophoresis.
How do differences in molecule size influence migration speed in a gel?
Smaller molecules travel faster through the gel pores, while larger molecules are slowed down and move more slowly.
What causes charged biomolecules to move through the gel during electrophoresis?
Charged biomolecules migrate because an electric current causes them to move toward the electrode with the opposite charge.
What is the basic setup for loading samples in gel electrophoresis?
Samples are loaded into wells within the gel and then subjected to an electric field to separate the molecules by size and charge.
How does a gel matrix help in separating different DNA fragments?
The gel matrix acts like a sieve, selectively impeding larger fragments more than smaller ones, resulting in distinct separation by length.
What is the role of the gel matrix in gel electrophoresis?
The gel matrix provides a porous medium through which molecules migrate at different speeds, allowing size-based separation.
Which gels are commonly used for separating large versus small molecules?
Agarose gels are typically used for large DNA molecules, while polyacrylamide gels are better suited for smaller molecules like RNA and small proteins.
Why is the electrophoresis chamber crucial for gel electrophoresis?
The electrophoresis chamber holds the gel and electrodes, creating the electric field that drives charged molecules toward the opposite electrode.
How are samples usually introduced into the gel?
Samples are loaded into wells at one end of the gel, often mixed with a tracking dye to visualize their migration.
Why is a buffer solution necessary in gel electrophoresis?
The buffer maintains the pH, provides ions for current flow, and dissipates heat generated during electrophoresis.
Why is the gel stained after electrophoresis?
The gel is stained so the separated molecules, like DNA, become visible for subsequent analysis.
How do ethidium bromide and SYBR Green function in DNA visualization?
Ethidium bromide and SYBR Green intercalate with DNA, fluorescing under ultraviolet light to reveal the DNA bands.
What information can be obtained by analyzing band patterns in a stained gel?
Band patterns reveal the size, quantity, and purity of separated molecules for accurate assessment.
