Amplified Fragment Length Polymorphism (AFLP)

Amplified Fragment Length Polymorphisms (AFLPs) are differences in restriction fragment lengths caused by SNPs or INDELs that create or abolish restriction endonuclease recognition sites.

The AFLP technique is based on the selective PCR amplification of restriction fragments from a total digest of genomic DNA.

After final amplification, selectively amplified fragments are separated by gel electrophoresis and visualized autoradiographically. MseI-MseI fragments are excluded from the autorad because only EcoRI-directed primers are normally labeled. Typically, the autorad has 100-300 fingerprints with sizes ranging from 80 to 500 nucleotides. Only a subset (10-40) of these total bands is polymorphic between two related individuals, such as Arabidopsis thaliana Columbia and Landsberg erecta ecotypes.

Using 3-bp selective primer extensions gives 128 possible linker combinations. Therefore, 128 subsets of genomic DNA can be readily amplified. Thus, thousands of markers can be generated quite rapidly.

Weaknesses of AFLP

l        Proprietary technology is needed to score heterozygotes and ++ homozygotes. Otherwise, AFLP must be dominantly scored.

l        Developing locus-specific markers from individual fragments can be difficult.

l        Need to use different kits adapted to the size of the genome being analyzed.