Difference Between RAPD and RFLP

The main difference between RAPD and RFLP is that RAPD is a type of PCR which amplifies random fragments of DNA in a large template by using short primers whereas, in RFLP, one or more restriction enzymes digest the DNA sample, producing restriction fragments then separated by gel electrophoresis. Therefore, RADP only requires a small amount of DNA for the assay while RFLP requires a large amount of DNA. Furthermore, RAPD is a rapid process, which can detect 1-10 loci while RFLP is a slower process, which can only detect 1-3 loci. In addition, RAPD is unable to detect allelic variants while RFLP can detect allelic variants. 

RAPD (random amplified polymorphic DNA) and RFLP (restriction fragment length polymorphism) are two techniques in molecular biology to detect genetic markers, which are short DNA sequences, exploiting variations in homologous DNA. 

Key Areas Covered 

1. What is RAPD
     – Definition, Procedure, Importance
2. What is RFLP
     – Definition, Procedure, Importance
3. What are the Similarities Between RAPD and RFLP
     – Outline of Common Features
4. What is the Difference Between RAPD and RFLP
     – Comparison of Key Differences

Key Terms 

Genetic Markers, PCR, RAPD, Restriction Digestion, RFLP 

What is RAPD 

RAPD or random amplified polymorphic DNA is a fast, PCR-based method for the detection of DNA variation. Two laboratories (Williams et. al., 1990; Welsh and McClelland, 1990) independently developed the technique. Moreover, RAPD uses a single, arbitrary primer for the amplification of many discrete DNA products. The steps involved in RAPD are as follows: 

1. DNA extraction 
2. Amplification by random primers 
3. Gel electrophoresis and  visualization of markers
   

   Figure 1: Lettuce Diversity by RAPD Markers

Furthermore, by resolving the resulting patterns, a semi-unique DNA profile of a particular DNA sample can be generated. Due to the less specificity of the random primer used in the assay, comparatively a higher number of loci can be amplified per random primer. However, RAPD is an important technique for gene mapping, population genetics, molecular evolutionary genetics, animal and plant breeding, etc. Moreover, it is vital for beginners by means of speed, cost, efficiency as well as the number of markers generated. 

What is RFLP 

RFLP or restriction length polymorphism is a method which exploits variation in homologous DNA. It is also a well-known method for its discrimination power. However, since RFLP is a non-PCR-based method, it uses restriction enzymes for the generation of fragments of the genome. Furthermore, RFLP detects the variations of lengths of restriction fragments among individuals. The three steps involved in RFLP are as follows: 

1. Restriction digestion of DNA 
2. Gel electrophoresis 
3. Southern blotting by specific  probes and detection
   

   Figure 2: Detection of Allelic Variants by RFLP

Moreover, restriction enzymes used in the process are unique for a particular DNA sequence. When the recognition sequence of the restriction enzymes is shorter, a greater number of fragments can be generated. Moreover, RFLP is important in genotyping, forensics, paternity testing, the detection of patterns in hereditary diseases, and in the detection of disease carriers. However, it is a labour-intensive and time-consuming process, which requires a large amount of DNA. 

Similarities Between RAPD and RFLP  

• RAPD and RFLP are two techniques in molecular biology to detect genetic markers, which exploit variation in homologous DNA. 
• They detect DNA polymorphism required for genetic mapping, genome fingerprinting, and for the investigation of genetic relatedness. 
• Therefore, they help to distinguish individuals, species or populations. 
• Moreover, both methods use the total amount of  DNA in the genome for the analysis.   

Difference Between RAPD and RFLP 

Definition 

RAPD (random amplified polymorphic DNA) refers to a PCR-based technique for identifying genetic variation while RFLP (restriction fragment length polymorphism) refers to a molecular method of genetic analysis, which allows individuals to be identified based on unique patterns of restriction enzyme cutting in specific regions of DNA.  

Type of method 

Moreover, RAPD is a PCR-based method while RFLP is a non-PCR-based method. 

Method 

While RAPD involves the amplification of genetic markers in a genome by using random primers, RFLP involves the restriction digestion of the genomic DNA. 

DNA Quantity 

In addition, RAPD requires a small quantity of DNA (10-50 ng) for the analysis while RFLP requires comparatively a large quantity of DNA (2-10 μg). 

Specificity for Species 

RAPD uses random primers universal for any species while RFLP uses different species-specific probes. 

Efficiency  

Furthermore, RAPD has fewer steps and it is a rapid process while RFLP has more steps as it is a slow process. 

Reliability 

RAPD is less reliable while RFLP is more reliable. 

Detection Capacity 

Another difference between RAPD and RFLP is that RAPD can detect 1-10 loci while RFLP can only detect 1-3 loci. 

Detection of Allelic Variants 

Furthermore, RAPD is unable to detect allelic variants while RFLP can detect allelic variants. 

Importance

RAPD is an important technique for gene mapping, population genetics, molecular evolutionary genetics, animal and plant breeding, etc. while RFLP is important in genotyping, forensics, paternity testing, the detection of patterns in hereditary diseases, and in the detection of disease carriers.

Conclusion 

RAPD is a PCR-based technique to detect genetic markers present in a genome. Moreover, it uses random primers for the amplification of these markers. As RAPD is a PCR-based technique, it only requires a small amount of DNA. In addition, it is a fast method but, it is unable to detect allelic variants. In contrast, RFLP is a non-PCR-based method which detects DNA variation within a genome. However, it uses restriction digestion of DNA, Southern blotting and detection of sequences by using specific probes. Therefore, it is a more reliable method. But, it requires a large amount of DNA. Therefore, the main difference between RAPD and RFLP is procedure and importance. 

References:

1. “Random Amplified Polymorphic DNA (RAPD).” National Center for Biotechnology Information, U.S. National Library of Medicine, Available Here.
2. “Restriction Fragment Length Polymorphism (RFLP).” National Center for Biotechnology Information, U.S. National Library of Medicine, Available Here.

Image Courtesy:

1. “RAPD” By Llull~commonswiki assumed – Own work assumed (Public Domain) via Commons Wikimedia  
2. “RFLP-figure” By Kenneth.jh.han – Own work (CC BY-SA 3.0) via Commons Wikimedia