Gel ElectrophoresisGel Electrophoresis

Electrophoresis: DNA Electrophoresis: DNA SeparationSeparation

Standard tool in Standard tool in biochemistry labsbiochemistry labs

UsesUses Diagnose diseaseDiagnose disease Identify genes and gene Identify genes and gene

structuresstructures Human genome projectHuman genome project Understand evolution of Understand evolution of

plants and animalsplants and animals Genetic engineering of Genetic engineering of

organisms (Example: organisms (Example: drought resistant cropsdrought resistant crops

Forensic scienceForensic science

DNA!DNA!

Extracted from animal, plant, and Extracted from animal, plant, and bacteria cellsbacteria cells

Individual cells are split open, and the Individual cells are split open, and the DNA is separated from the rest of the DNA is separated from the rest of the cellular debriscellular debris

DNA is then treated with special proteins DNA is then treated with special proteins called restriction enzymes, which cleave called restriction enzymes, which cleave the DNA into smaller fragmentsthe DNA into smaller fragments

How does Electrophoresis How does Electrophoresis work?work?

DNA molecules are DNA molecules are negatively charged negatively charged

Use electricity to Use electricity to separate DNA separate DNA protein molecules protein molecules based on charge and based on charge and massmass

DNA samples are DNA samples are taken from animal or taken from animal or plant cellsplant cells

Agarose GelAgarose Gel

Used as the support Used as the support material to separate material to separate DNA moleculesDNA molecules

Derived from Derived from seaweedseaweed

Note “wells”- DNA Note “wells”- DNA solution is loaded solution is loaded into these holesinto these holes

Loading the GelLoading the Gel

DNA loaded into gel: DNA loaded into gel: mixture of different mixture of different sized DNA fragments sized DNA fragments

Loading the GelLoading the Gel

Loading gel with DNA mixture + dyeLoading gel with DNA mixture + dye Gel is suspended in buffer which conducts electrical currentGel is suspended in buffer which conducts electrical current

Separation of DNASeparation of DNA

Note applied electrical Note applied electrical charge- DNA is charge- DNA is negatively charged and negatively charged and will migrate to the will migrate to the positive polepositive pole

Gel matrix acts as a Gel matrix acts as a “seive” for DNA“seive” for DNA

Large DNA molecules Large DNA molecules cannot pass through the cannot pass through the small holes in the gelsmall holes in the gel

Small molecules move Small molecules move easily through the geleasily through the gel

Running the GelRunning the Gel

Electric current is applied to gelElectric current is applied to gel DNA starts to migrate through the gelDNA starts to migrate through the gel

Separation of DNASeparation of DNA

As separation As separation continues, the continues, the smaller fragments smaller fragments move farther down move farther down the gelthe gel

Molecular MarkersMolecular Markers

A DNA molecular A DNA molecular marker “ladder” is marker “ladder” is run at the same time run at the same time as your sample DNAas your sample DNA

Markers are of Markers are of known molecular known molecular weightsweights

Markers used to Markers used to estimate the sizes of estimate the sizes of your sample DNAyour sample DNA

Reading the GelReading the Gel

Dye in gel reacts with Dye in gel reacts with UV light, DNA is UV light, DNA is fluorescentfluorescent

Photo takenPhoto taken