7/24/2019 Gel Electrophoresis of Nucleic Acids

1/11

ELIYAH A. TABILOG

Gel electrophoresisof nucleic acids

7/24/2019 Gel Electrophoresis of Nucleic Acids

2/11

is an analytical technique used to separate DNA or RNA fragmby size and reactivity. Nucleic acid molecules which are to be anare set upon a viscous medium, the gel, where an electric field the nucleic acids to migrate toward the anode, due to the net necharge of the sugar-phosphate backbone of the nucleic acid cha

WHAT is

Nucleic acid electrophoresis?

7/24/2019 Gel Electrophoresis of Nucleic Acids

3/11

Nucleic acidsare biopolymers, or large biomolecules, essentall known forms of life. Nucleic acids, whichinclude DNA (deoxyribonucleic acid) and RNA (ribonucleicare made from monomers known as nucleotides.

WHATIS NUCLEIC ACID

7/24/2019 Gel Electrophoresis of Nucleic Acids

4/11

How fast will the DNA migrate?

+-

Power

DNA

smalllarge

7/24/2019 Gel Electrophoresis of Nucleic Acids

5/11

Size of DNA

Conformation of DNA

Concentration of ethidium bromide

Gel concentration

Applied Electric field

actors a!ecti"g migratio" of"ucleic acids

7/24/2019 Gel Electrophoresis of Nucleic Acids

6/11

Agarose gel electrophoresis

is a method of gel electrophoresis used in biochemistry, mbiology, genetics, and clinical chemistry to separate apopulation of DNA or proteins in a matrix of agarose.

Pulsed field gel electrophoresis

is a technique used for the separation of large deoxyriboacid (DNA) molecules by applying to a gel matrix an electrthat periodically changes direction.

Si#e of DNA

7/24/2019 Gel Electrophoresis of Nucleic Acids

7/11

The conformation of the DNA molecule can significantly affemovement of the DNA.

Supercoiled DNA moves faster

Relaxed DNA move slow

Co"formatio" of DNA

7/24/2019 Gel Electrophoresis of Nucleic Acids

8/11

Circular DNA are more strongly affected by ethidium bromidconcentration than linear DNA if ethidium bromide is presengel during electrophoresis.

Increasing ethidium bromide intercalated into the DNA canit from a negatively supercoiled molecule into a fully relaxed then to positively coiled superhelix at maximum intercalation

Agarose gel electrophoresis can be used to resolve circular Dwith different supercoiling topology.

Co"ce"tratio" of Ethidium $rom

7/24/2019 Gel Electrophoresis of Nucleic Acids

9/11

The concentration of the gel determines the pore size of the gwhich affect the migration of DNA.

Increasing the agarose concentration of a gel reduces the migspeed and improves separation of smaller DNA molecules,

%el co"ce"tratio"

7/24/2019 Gel Electrophoresis of Nucleic Acids

10/11

At low voltages, the rate of migration of the DNA is proportithe voltage applied.

For optimal resolution of DNA > 2kb in size in standard gelelectrophoresis, 5 to 8 V/cm is recommended.

Applied Electric &eld

7/24/2019 Gel Electrophoresis of Nucleic Acids

11/11