AFLP-PCR or just AFLP is a PCR-based tool used in

genetics Genetics is the study of genes, genetic variation, and heredity in organisms.Hartl D, Jones E (2005) It is an important branch in biology because heredity is vital to organisms' evolution. Gregor Mendel, a Moravian Augustinian friar wor ...

research,

DNA fingerprint DNA profiling (also called DNA fingerprinting) is the process of determining an individual's DNA characteristics. DNA analysis intended to identify a species, rather than an individual, is called DNA barcoding. DNA profiling is a forensic tec ...

ing, and in the practice of

genetic engineering Genetic engineering, also called genetic modification or genetic manipulation, is the modification and manipulation of an organism's genes using technology. It is a set of technologies used to change the genetic makeup of cells, including t ...

. Developed in the early 1990s by KeyGene, AFLP uses

restriction enzymes A restriction enzyme, restriction endonuclease, REase, ENase or'' restrictase '' is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class o ...

to digest

genomic DNA Genomic deoxyribonucleic acid (abbreviated as gDNA) is chromosomal DNA, in contrast to extra-chromosomal DNAs like plasmids. Most organisms have the same genomic DNA in every cell; however, only certain genes are active in each cell to allow for ...

, followed by

ligation Ligation may refer to: * Ligation (molecular biology), the covalent linking of two ends of DNA or RNA molecules * In medicine, the making of a ligature (tie) * Chemical ligation, the production of peptides from amino acids * Tubal ligation, a meth ...

of adaptors to the

sticky ends DNA ends refer to the properties of the ends of linear DNA molecules, which in molecular biology are described as "sticky" or "blunt" based on the shape of the complementary strands at the terminus. In sticky ends, one strand is longer than the ...

of the restriction fragments. A subset of the restriction fragments is then selected to be amplified. This selection is achieved by using

primer Primer may refer to: Arts, entertainment, and media Films * ''Primer'' (film), a 2004 feature film written and directed by Shane Carruth * ''Primer'' (video), a documentary about the funk band Living Colour Literature * Primer (textbook), a t ...

s complementary to the adaptor sequence, the restriction site sequence and a few nucleotides inside the restriction site fragments (as described in detail below). The amplified fragments are separated and visualized on denaturing on

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 ...

, either through autoradiography or

fluorescence Fluorescence is the emission of light by a substance that has absorbed light or other electromagnetic radiation. It is a form of luminescence. In most cases, the emitted light has a longer wavelength, and therefore a lower photon energy, tha ...

methodologies, or via automated capillary sequencing instruments. Although AFLP should not be used as an acronym, it is commonly referred to as "Amplified fragment length polymorphism". However, the resulting data are not scored as length polymorphisms, but instead as presence-absence polymorphisms. AFLP-PCR is a highly sensitive method for detecting polymorphisms in DNA. The technique was originally described by Vos and Zabeau in 1993. In detail, the procedure of this technique is divided into three steps: #Digestion of total cellular DNA with one or more

restriction enzyme A restriction enzyme, restriction endonuclease, REase, ENase or'' restrictase '' is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class o ...

s and ligation of restriction half-site specific adaptors to all restriction fragments. #Selective amplification of some of these fragments with two PCR primers that have corresponding adaptor and restriction site specific sequences. # Electrophoretic separation of

amplicon In molecular biology, an amplicon is a piece of DNA or RNA that is the source and/or product of amplification (molecular biology), amplification or DNA replication, replication events. It can be formed artificially, using various methods including ...

s on a gel matrix, followed by visualisation of the band pattern.

Applications

The AFLP technology has the capability to detect various polymorphisms in different genomic regions simultaneously. It is also highly sensitive and reproducible. As a result, AFLP has become widely used for the identification of genetic variation in strains or closely related species of plants, fungi, animals, and bacteria. The AFLP technology has been used in criminal and paternity tests, also to determine slight differences within populations, and in linkage studies to generate maps for

quantitative trait locus A quantitative trait locus (QTL) is a locus (section of DNA) that correlates with variation of a quantitative trait in the phenotype of a population of organisms. QTLs are mapped by identifying which molecular markers (such as SNPs or AFLPs) c ...

(QTL) analysis. There are many advantages to AFLP when compared to other marker technologies including randomly amplified polymorphic DNA ( RAPD), restriction fragment length polymorphism (RFLP), and

microsatellite A microsatellite is a tract of repetitive DNA in which certain DNA motifs (ranging in length from one to six or more base pairs) are repeated, typically 5–50 times. Microsatellites occur at thousands of locations within an organism's genome. ...

s. AFLP not only has higher reproducibility, resolution, and sensitivity at the whole genome level compared to other techniques, but it also has the capability to amplify between 50 and 100 fragments at one time. In addition, no prior sequence information is needed for amplification (Meudt & Clarke 2007). As a result, AFLP has become extremely beneficial in the study of taxa including bacteria, fungi, and plants, where much is still unknown about the genomic makeup of various organisms. The AFLP technology is covered by patents and patent applications of Keygene N.V. AFLP is a registered trademark of Keygene N.V.

References

External links

Software for analyzing AFLP data
BioNumerics
One universal platform to manage and analyze all your biological data including AFLP
KeyGene Quantar Suite
''Versatile marker scoring software''
SoftGenetics
''GeneMarker'' fragment analysis software Freeware for analyzing AFLP data
SourceForge
''Genographer'' Free software for manual scoring (Java application)
SourceForge
''RawGeno'' Free automated scoring (R CRAN environment, including a user-friendly GUI) Online programs for simulation of AFLP-PCR

- BProkaryotes or uploaded sequences *In silico AFLP-PCR fo
prokaryotes
som
eukaryotes
o
uploaded sequences
*Enzymes for AFL
New England Biolabs

AFLP Technology note at KeyGene

AFLP Applications
{{DEFAULTSORT:Amplified Fragment Length Polymorphism Molecular biology DNA DNA profiling techniques