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RFLP (Restriction Fragment Length Polymorphism)

Restriction fragment length polymorphism (RFLP) is considered to be the simplest and earliest method to detect SNPs. SNP-RFLP makes use of the many different restriction endonucleases and their high affinity to unique and specific restriction sites. By performing a digestion on a genomic sample and determining fragment lengths through a gel assay it is possible to ascertain whether or not the enzymes cut the expected restriction sites. A failure to cut the genomic sample results in an identifiably larger than expected fragment implying that there is a mutation at the point of the restriction site which is rendering it protection from nuclease activity.

Unfortunately, the combined factors of the high complexity of most eukaryotic genomes, the requirement for specific endonucleases, the fact that the exact mutation cannot necessarily be resolved in a single experiment and the slow nature of gel assays make RFLP a poor choice for high throughput analysis.