Difference Between AFLP and RFLP

Key Difference – AFLP vs RFLP
 

DNA studies have an immense importance in understanding and determining phylogenetic relationships, diagnosing genetic diseases and mapping organism’s genomes. Several techniques associated with DNA analysis are also used for the identification of a particular gene or a DNA sequence in a pool of unknown DNA. They are known as molecular markers. Amplified Fragment Length Polymorphism (AFLP) and Restriction Fragment Length Polymorphism (RFLP) are two such molecular markers (methods) developed in molecular biology to detect genetic variation between organisms. Both methods are equally important and have advantages and disadvantages. The key difference between AFLP and RFLP is that AFLP involves selective PCR amplification of the digested DNA while RFLP does not involve selective PCR amplification of the DNA fragments.

CONTENTS
1. Overview and Key Difference
2. What is AFLP
3. What is RFLP
4. Side by Side Comparison – AFLP vs RFLP
5. Summary

What is AFLP?

AFLP (Amplified Fragment Length Polymorphism) is an important tool in molecular biology and is extensively used in genetic variation analysis. AFLP is based on the specific PCR amplification of the fragmented genomic DNA and detection of the polymorphism by autoradiographs via gel electrophoresis. AFLP widely contributes to identifying genetic differences in strains or closely related species of various kingdoms including plants, animals, bacteria and fungi. AFLP can be carried out with small amounts of unknown DNA samples. It does not require prior sequence knowledge and designing of probes.

Steps of AFLP

1. Isolation of DNA
2. Digestion of the DNA with restriction endonucleases
3. Ligation of the restricted DNA fragments with adaptors
4. Selective amplification of the fragments with specific restriction sites
5. Separation of the PCR products by gel electrophoresis
6. Visualization of the gel matrix by autoradiograph

AFLP is a more sensitive and reproducible method which can be used in DNA profiling of several taxa including fungi, bacteria, plants and animal without prior knowledge of DNA sequences. It helps in identifying slight differences among individuals in populations due to its highly sensitive nature. AFLP is also important in genome mapping, forensic studies, parental testing, genotyping, etc.

What is RFLP?

Restriction Fragment Length Polymorphisms (RFLPs) is a technique that is used to detect the genetic variations in homologous DNA sequences. It is the first method developed for DNA profiling. Organisms have unique DNA fingerprints or DNA profiles. RFLP serves as an important tool to analyze the variation between DNA profiles of intraspecific or closely related organisms since homologous sequences have different restriction sites (locations) that are unique to a particular organism. When homologous DNA are digested with specific restriction endonucleases, it will result in different DNA profiles which are unique to each individual. Therefore, the principal of this method is the detection of genetic variation among organisms by restricting homologous DNA with specific restriction enzymes and analysis of the fragment length polymorphism via gel electrophoresis and blotting. Blotting patterns are unique to each organism and characterize the specific genotypes.

Steps of RFLP

1. Isolation of sufficient amount of DNA from samples
2. Fragmentation of the DNA samples with specific restriction endonucleases into short sequence
3. Separation of the resulted fragments with different lengths by agarose gel electrophoresis.
4. Transfer of the gel profile into a membrane by Southern blotting

5. Hybridization of the membrane with labeled probes and analysis of the fragment length polymorphism in each profile

RFLP is a very important technique in detecting disease inheritance and finding the risk of the disease occurrence among family members. RFLP is also frequently used in genome mapping, identifying criminals in forensics, paternity testing, etc. RFLP has several limitations as well. RFLP requires the prior knowledge of sequence data to design probes for hybridization. It also requires isolation of sufficient amount of DNA from the sample to analyze, which is difficult in forensic studies.

What is the difference between AFLP and RFLP?

ALFP vs RFLP
AFLP involves selective PCR amplification of the digested DNA.	RFLP does not involve PCR unless it is PCR-RFLP.
Sequence Knowledge
Prior sequence knowledge is not required.	Prior sequence knowledge is required to designed RFLP probes.
Reliability
This is more reliable.	This is less reliable compared to AFLP.
Efficiency in Detecting Polymorphism
This has a higher efficiency in detecting polymorphism than RFLP.	This is less efficient compared to AFLP.
Cost
This is a little expensive compared to RFLP.	This is less expensive compared to AFLP.
Applications
AFLPs have been applied to genome mapping, DNA fingerprinting, genetic diversity studies, paternity testing and forensics	RFLP analysis is an important tool in genome mapping, localization of genes for genetic disorders, determination of risk for disease, and paternity testing.

Summary – AFLP vs RFLP

AFLP and RFLP are two techniques used as genetic markers for the evaluation of diversity and assessment of the genetic relationships in molecular biology. AFLP serves as an efficient and sensitive method for detection of genetic polymorphism between organisms than RFLP. However, though both these methods have different efficiencies in detecting genetic variations, they are still used for DNA fingerprinting and disease diagnosis.

Reference:
`1. Garcia, Antonio A. F., Luciana L. Benchimol, Antônia M. M. Barbosa, Isaias O. Geraldi, Souza Jr. Cláudio L., and Anete P. De Souza. “Comparison of RAPD, RFLP, AFLP and SSR markers for diversity studies in tropical maize inbred lines.” Genetics and Molecular Biology. Sociedade Brasileira de Genética, 2004. Web. 19 Mar. 2017
2. “Restriction Fragment Length Polymorphism (RFLP).” National Center for Biotechnology Information. U.S. National Library of Medicine, n.d. Web. 19 Mar. 2017
3. Masiga D. K. and Turner C. M. (2004). “Amplified (restriction) fragment length polymorphism (AFLP) analysis”. Methods Mol Biol: 270: 173-86. NCBI. Web. 19 Mar. 2017

Image Courtesy:
1. “ALFP” By Barbarossa at Dutch Wikipedia – Transferred from nl.wikipedia to Commons. (Public Domain) via Commons Wikimedia
2. “RFLP mapping” By Retama – Own work (GFDL) via Commons Wikimedia