[LAB] UNIT 4.3 Gel Electrophoresis Flashcards
This is the process of separating the charged
constituents of a sample by means of an electrical
current.
Electrophoresis
In electrophoresis, separation is done by means of _________ ________
electric current
2 main components of electrophoresis
Chamber
Power Source
Positively charged pole that attracts negative particles
Anode
Color of Anode
Red
Negatively charged pole that attracts positive particles
Cathode
Color of Cathode
Black
Flow of electric current
Cathode –> Anode –> Power Source
Electrophoresis is used to separate __________
charged particles.
negatively
An ion that contains two functional groups
Zwitterion
T or F: A zwitterion is an ion possessing both positive and negative electrical charges
T
T or F: A zwitterion is mostly positively charged
F
It is mostly neutral
T or F: DNA is negatively charged and will travel towards the cathode
F
It will travel towards the anode (positive pole)
Specific buffer needed for electricity to flow
Tris-Acetate-EDTA Buffer
Migration of charged macromolecules in the
presence of an electrical power through a porous
support
Zone Electrophoresis
T or F: Zone electrophoresis requires electrical power
T
Examples of porous support used in zone electrophoresis
Paper
Cellulose acetate
Agarose gel
Has a net charge that can be either positive or
negative depending on pH conditions
Amphoteric
Movement of buffer ions & solvent relative to the
fixed support
Electroendosmosis / Endosmosis
Migration of small charged ions
Iontophoresis
5 components of electrophoresis
- The driving force
- The support medium
- The buffer
- The sample
- The detecting system
Examples of general support media
Cellulose acetate
Agarose gel
Polyacrylamide gel
Carcinogenic stain used for visualization of analyte
Ethidium bromide (EtBr)
In the first step of electrophoresis, EtBr is added to a flask containing ________ _____
Agarose gel
T or F: Adding EtBr to agarose gel requires shaking or swirling
T
Added in order to track the
migration of the analyte across the gel
Tracking dye
Added by some manufacturers in their tracking dyes
Glycerol
T or F: DNA can be seen even in the absence of dyes
F
Example of a tracking dye
Cotton Phenol Blue
T or F: GelRed is an alternative fluorescent detection dye
T
T or F: EtBr is a detection dye
T
What equipment is used to place the analyte into the wells?
Pipette
This will give proteins the net electric (negative) charge that allows them to move from the negatively charged electrode to the positively charged electrode
Buffer
Red wire provides a _______ charged electrode
Black wire provides the _______ charged
electrode
positively ; negatively
A standard piece of equipment used in life science laboratories for visualization of target DNA
UV Transilluminator
Unit used in electrophoresis
bp (base pairs)
It is a reference to estimate the size of unknown DNA molecules that were separated
DNA Ladder
T or F: The closer the sample to the well, the heavier it is. The farther the sample to the well, the lighter it is.
T
The best stain to use for DNA extraction is _______ _______ because it allows for intercalation or insertion between layers
ethidium bromide
T or F: Gel M is safer and more sensitive than EtBr
F
It is less sensitive
T or F: DNA Ladder is treated the same as the samples
T
T or F: Turn on the power prior to running
T
T or F: Results of electrophoresis is merely quantitative
F
Results are both quantitative and qualitative
Fractions of Separated Proteins
- Albumin
- Alpha 1
- Alpha 2
- Beta
- Gamma globulins
Stains for visualization of bands
● Amido Black
● Ponceau S
● Oil Red O
● Sudan Black
● Fat Red 7B
● Coomassie Blue
● Gold/Silver Stain
The movement of molecules towards the gel is dependent of their
molecular weight
T or F: Lower voltages are used for longer periods of time, while higher voltages for shorter periods of time
T
T or F: The dye somewhat serves as the standard curve
F
The DNA Ladder somewhat serves as the standard curve
