What is RAPD primer?

Randomly amplified polymorphic DNA (RAPD) is a PCR-based technique which uses arbitrary primers which bind to the nonspecific sites on the DNA and amplify the DNA. These amplified fragments are then migrated on agarose gel and difference in the band pattern is observed.

How many primers are used in RAPD?

Unlike traditional PCR analysis, RAPD (pronounced “rapid”) does not require any specific knowledge of the DNA sequence of the target organism: the identical 10-mer primers will or will not amplify a segment of DNA, depending on positions that are complementary to the primers’ sequence.

What is the difference between PCR and RAPD?

RAPD stands for Random Amplification of Polymorphic DNA. RAPD reactions are PCR reactions, but they amplify segments of DNA which are essentially unknown to the scientist (random). Often, PCR is used to amplify a known sequence of DNA. Thus, PCR leads to the amplification of a particular segment of DNA.

What is needed in RAPD?

RAPD analyses generally require purified, high molecular weight DNA, and precautions are needed to avoid contamination of DNA samples because short random primers are used that are able to amplify DNA fragments in a variety of organisms.

What is primer in PCR?

A primer is a short, single-stranded DNA sequence used in the polymerase chain reaction (PCR) technique. In the PCR method, a pair of primers is used to hybridize with the sample DNA and define the region of the DNA that will be amplified. Primers are also referred to as oligonucleotides.

What is the difference between RFLP and RAPD?

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.

What is the full form of AFLP?

Amplified fragment length polymorphism (AFLP) is a PCR-based technique that uses selective amplification of a subset of digested DNA fragments to generate and compare unique fingerprints for genomes of interest.

What is Rapd PCR used for?

Random amplified polymorphic DNA (RAPD) is a PCR based technique for identifying genetic variation. It involves the use of a single arbitrary primer in a PCR reaction, resulting in the amplification of many discrete DNA products. The technique was developed independently by two different laboratories (Williams et.

What are the applications of RAPD?

As an extension to the variety of existing techniques using polymorphic DNA markers, the Random Amplified Polymorphic DNA (RAPD) technique may be used in molecular ecology to determine taxonomic identity, assess kinship relationships, analyse mixed genome samples, and create specific probes.

Why are 2 primers needed for PCR?

Two primers are used in each PCR reaction, and they are designed so that they flank the target region (region that should be copied). That is, they are given sequences that will make them bind to opposite strands of the template DNA, just at the edges of the region to be copied.

What makes a good primer for PCR?

A good length for PCR primers is generally around 18-30 bases. The shorter the primers are, the more efficiently they will bind or anneal to the target. Try to make the melting temperature (Tm) of the primers between 65°C and 75°C, and within 5°C of each other.

What is the principle of RFLP?

RFLP is one of the earliest molecular markers developed for genetic mapping. The principle of RFLP markers is that any genomic DNA can be differentiated according to the presence or absence of restriction enzyme sites. Restriction enzymes recognize and cut at the particular site.

How is the RAPD protocol optimized for PCR?

RAPD protocol was optimized based on the use of lower concentrations of primer (2 μM) and Taq polymerase (2 units), 50 ng of template DNA, higher concentration of MgCl 2 (2 mM) and an annealing temperature of 37°C, resulted optimal amplification.

What kind of PCR buffer is used for RAPD?

Each 15 μL reaction volume contained about 50 ng of template DNA, 1X PCR Buffer (10 mM Tris HCl pH 8; 50 mM KCl), 3 mM MgCl 2 (Sigma, USA), 0.2 mM dNTP Mix, 0.5 μM of single primer, 0.2 U of Taq DNA polymerase (Helini Biomolecules, India).

How is PCR used to isolate RAPD DNA?

PCR products were electrophoresed on 2% (w/v) agarose gels, in 1X TBE Buffer at 50 V for 3 h and then stained with ethidiumbromide (0.5 μg mL -1 ). Gels with amplification fragments were visualized and photographed under UV light.

How much DNA is needed for RAPD analysis?

The yield of DNA ranged from 1.5-2.5 μg μL -1 per gram of the leaf tissue. The technique is ideal for isolation of DNA from different salt marsh plant species and used for Randomly Amplified Polymorphic DNA (RAPD) analysis.