Amplified Fragment Length Polymorphism
AFLP is a DNA fingerprinting technique that can be used to answer a variety of research questions. This PCR based tool utilizes restriction enzymes to digest genomic DNA to smaller fragments and consists of three main steps. The first step involves restriction digestion of DNA with two types of enzymes (a rare and a frequent cutter), followed by the ligation of double-stranded oligonucleotide adapters, specific to the ends of the generated fragments. There are several reasons for the use of two types restriction enzymes: generation of short DNA segments with good amplification and separation on the gel, the rare cutter will complement the frequent cutter by decreasing the number of generated fragments, and only one strand of the double stranded PCR products can be labeled. The second step consists of the amplification of certain restriction fragments with PCR primers (the primers are designed to target the adapters fused to the end of the fragments). And the third step is the analysis of the amplified fragments after visualization on a denaturing polyacrylamide gel.
AFLP is a powerful molecular tool allowing simultaneous screening of a large number of DNA regions that are spread randomly throughout the genome. One of the key features of AFLP analysis is the detection of polymorphisms of genomic DNA fragments obtained through restriction digestion. Further more, AFLP markers are very useful for studying genetic variation between closely related species. “For a wide range of taxa, including plants, fungi, animals and bacteria, AFLP markers have been used to uncover cryptic genetic variation of strains, or closely related species, that had been impossible to resolve with morphological or other molecular systematic characters” (Mueller et al). Besides detecting fragment polymorphisms, “The AFLP technique may be used for fingerprinting of cloned DNA segments like cosmids, P1 clones, bacterial artificial chromosomes or yeast artificial chromosomes (Vos et al). AFLP can also be applied to criminal investigations and paternity testing.
In addition, AFLP can be useful for studying genetic variation in different yeast species used in wine making, “Amplified fragment length polymorphism (AFLP) was used to investigate genetic variation in commercial strains, type strains and winery isolates from a number of yeast species” (Lopes et al). After performing a cluster analysis with the AFLP data obtained through this study, the genetic relatedness of Saccharomyces cerevisiae and Dekkera bruxellensis was determined. However, there are also limitations associated with this method, “The main limitation of AFLP in yeast systematics may be its inability to establish genetic similarities between species” (Lopes et al).
AFLP is very powerful molecular technique with a wide variety of applications in species identification, strain differentiation, and detection of genetic similarities between organisms within the same species. Therefore, AFLP can be used in a variety of evolutionary, phylogenetic, and linkage studies.
- Lopes, Miguel. "AFLP fingerprinting for analysis of yeast genetic variation." International Journal of Systematic Bacteriology49 (1999): 915-24. Print.
- Mueller, Ulrich G. "AFLP Genotyping and Fingerprinting." Trends in Ecology & Evolution 14 (1999): 384-84. Print.
- Vos, Pieter. "AFLP: a new technique for DNA fingerprinting." Nucleic Acids Research 25 (1995): 4407-414. Print.