Pulsed-field gel electrophoresis (PFGE) is a technique used for the separation of large

DNA Deoxyribonucleic acid (; DNA) is a polymer composed of two polynucleotide chains that coil around each other to form a double helix. The polymer carries genetic instructions for the development, functioning, growth and reproduction of al ...

molecule A molecule is a group of two or more atoms that are held together by Force, attractive forces known as chemical bonds; depending on context, the term may or may not include ions that satisfy this criterion. In quantum physics, organic chemi ...

s by applying an

electric field An electric field (sometimes called E-field) is a field (physics), physical field that surrounds electrically charged particles such as electrons. In classical electromagnetism, the electric field of a single charge (or group of charges) descri ...

that periodically changes direction to a gel matrix. Unlike standard

agarose gel electrophoresis Agarose gel electrophoresis is a method of gel electrophoresis used in biochemistry, molecular biology, genetics, and clinical chemistry to separate a mixed population of macromolecules such as DNA or proteins in a matrix of agarose, one of the t ...

, which can separate DNA fragments of up to 50 kb, PFGE resolves fragments up to 10 Mb. This allows for the direct analysis of genomic DNA.

History

In 1984, David C. Schwartz and

Charles Cantor Charles R. Cantor (born August 26, 1942 in Brooklyn) is an American molecular geneticist who, in conjunction with David Schwartz, developed pulse field gel electrophoresis for very large DNA molecules. Cantor's three-volume book ''Biophysical ...

published the first successful application of alternating electric fields for the separation of large DNA molecules. This technique, which they named PFGE, resulted in the development of several variations, including Orthogonal Field Alternation Gel Electrophoresis (OFAGE), Transverse Alternating Field Electrophoresis (TAFE), Field-Inversion Gel Electrophoresis (FIGE), and Clamped Homogeneous Electric Fields (CHEF), among others.

Procedure

The procedure for PFGE is similar to that of standard

, with the main exception being the application of the electric current. Generally, in PFGE electrophoresis chambers, the voltage periodically switches between three directions: one along the central axis, and two at a 60 degree angle along each side. The application of the voltage can change depending on the variation of PFGE used.

Applications

PFGE may be used for

genotyping Genotyping is the process of determining differences in the genetic make-up (genotype) of an individual by examining the individual's DNA sequence using bioassay, biological assays and comparing it to another individual's sequence or a reference seq ...

genetic fingerprinting DNA profiling (also called DNA fingerprinting and genetic fingerprinting) is the process of determining an individual's deoxyribonucleic acid (DNA) characteristics. DNA analysis intended to identify a species, rather than an individual, is cal ...

. It has commonly been considered a

gold standard A gold standard is a backed currency, monetary system in which the standard economics, economic unit of account is based on a fixed quantity of gold. The gold standard was the basis for the international monetary system from the 1870s to the ...

epidemiological Epidemiology is the study and analysis of the distribution (who, when, and where), patterns and Risk factor (epidemiology), determinants of health and disease conditions in a defined population, and application of this knowledge to prevent dise ...

studies of pathogenic organisms for several decades. For instance, subtyping bacterial isolates with this method has made it easier to discriminate among strains of ''

Listeria monocytogenes ''Listeria monocytogenes'' is the species of pathogenic bacteria that causes the infection listeriosis. It is a facultative anaerobic bacterium, capable of surviving in the presence or absence of oxygen. It can grow and reproduce inside the ho ...

'', ''

Lactococcus garvieae ''Lactococcus garvieae'' is a known fish pathogen affecting saltwater fish in the Far East, specifically in rainbow trout, Japanese yellowtail, Cobia (''Rachycentron canadum'') and grey mullet (''Mugil cephalus''). This bacteria causes lesions ...

'' and some clinical isolates of ''

Bacillus cereus ''Bacillus cereus'' is a Gram-positive bacteria, Gram-positive Bacillus, rod-shaped bacterium commonly found in soil, food, and marine sponges. The specific name, ''cereus'', meaning "waxy" in Latin, refers to the appearance of colonies grown o ...

'' group isolated from diseases aquatic organisms and thus to link environmental or food isolates with clinical infections. It is now in the process of being superseded by next generation sequencing methods.

References

External links

Pulse field methodApplied Maths
''BioNumerics'' PFGE typing {{DEFAULTSORT:Pulsed Field Gel Electrophoresis Biological techniques and tools Electrophoresis Genetics techniques

History

Procedure

Applications

See also

References

External links