AFLP

AFLP, or Amplified Fragment Length Polymorphism, is a DNA fingerprinting technique used to detect genetic variation without requiring prior sequence information. Developed in 1995 by Vos and colleagues, AFLP combines restriction digestion of genomic DNA with selective PCR amplification to generate a large number of polymorphic fragments, enabling high-resolution profiling across the genome.

The method begins with the digestion of genomic DNA using two restriction enzymes, typically a rare cutter (such as PstI) and a frequent cutter (such as MseI). Adapters are ligated to the resulting ends to provide known primer binding sites. A pre-selective amplification uses primers complementary to the adapters and restriction sites, and a subsequent selective amplification adds extra nucleotides at the 3' end of the primers to increase selectivity. The amplified fragments are separated by gel electrophoresis or capillary electrophoresis and visualized; the presence or absence of bands at specific sizes is scored as binary data.

AFLP has broad applications in genetics and breeding. It is widely used for constructing genetic linkage maps, assessing genetic diversity and population structure, identifying cultivar or strain markers, and supporting quantitative trait locus (QTL) mapping. While AFLP generally provides higher genome coverage and discriminatory power than older marker systems such as RFLP and RAPD, it is a dominant marker, meaning it does not directly distinguish homozygous from heterozygous states. It also requires more specialized equipment and careful standardization to ensure reproducibility and comparability of results across laboratories.

Limitations include dependence on DNA quality, potential scoring ambiguity for faint or closely spaced bands, and reduced transferability of markers between different platforms or experiments. Despite these considerations, AFLP remains a robust tool for genome-wide polymorphism analysis in a variety of organisms.