Difference Between Forward and Reverse Primer

Key Difference – Forward vs Reverse Primer
 

Polymerase chain reaction (PCR) is a DNA amplification method that is used in Molecular Biological applications. It is a commonly used technique that makes millions to billions of copies of a particularly interested DNA sequence. It is an in vitro method performed in laboratories. PCR technique is totally dependent on the commercially produced DNA polymerase called Taq polymerase. And also it requires several other components and proper temperature maintenance. One important component is primers. Primers are the short DNA sequences specifically designed for the target DNA sequence. They are usually around 20 nucleotides in length. Taq polymerase catalyzes the adding of nucleotides into preexisting nucleotide sequence. Hence, primers are served as starting points of the synthesis of new strands. Taq polymerase works only in 5’ to 3’ direction hence the DNA synthesis occurs in the same 5’ to 3’ direction. Since DNA is double stranded, two types of primers are needed in PCR. They are known as forward primer and reverse primer. Forward and reverse primers are termed based on the direction of the elongation of the primer in DNA when DNA synthesis occurs. Forward primer anneals with the antisense DNA strand and initiates the synthesis of +ve strand of the gene into 5’ to 3’ direction. Reverse primer anneals with the sense strand and initiates the synthesis of the complementary strand of the coding strand; which is –ve the strand of the gene into 5’to 3’ direction. This is the key difference between forward and reverse primers.

CONTENTS

1. Overview and Key Difference
2. What is a Forward Primer
3. What is a Reverse Primer
4. Similarities Between Forward and Reverse Primer
5. Side by Side Comparison – Forward vs Reverse Primer in Tabular Form
6. Summary

What is a Forward Primer?

Forward orientation is the synthesis of the coding strand or the sense strand of a gene. Taq polymerase catalyzes the synthesis of a new strand in 5’ to 3’ direction. The synthesis of coding strand occurs when the primer anneals with the noncoding or the antisense strand and elongates in 5’ to 3’ direction.

The primer that anneals with the antisense strand or the noncoding strand or the template strand is known as forward primer since forward primer acts as a starting point to the synthesis of coding or the positive strand of the gene. Forward primer has a short nucleotide sequence that is complementary to the 3’ flanking end of the antisense strand. It hybridizes with the antisense strand and facilitates the Taq polymerase to add nucleotides that are complementary to the template strand.

What is a Reverse Primer?

Reverse primer is the short DNA sequence that anneals with the 3’ end of the sense strand or the coding strand. Reverse primer serves as the starting point to synthesize a complementary strand of the coding sequence or the noncoding sequence. Reverse primer is designed complementary to the 3’ end of the coding strand. Hence, it anneals with the flanking 3’ end of the coding strand and allows Taq polymerase to synthesize the antisense strand or the template strand. Since its orientation is in a reversed manner, this primer is labeled as reverse primer.

Both reverse and forward primers are important for the production of millions to billions of copies of particular regions of DNA that are targeted or interested.

What are the Similarities Between Forward and Reverse Primer?

• Both Forward and Reverse primers are made from oligonucleotides.
• Both Forward and Reverse Primers possess short nucleotide sequence complementary to the flanking ends of the DNA double strands.
• Both Forward and Reverse primers usually consist of 20 nucleotides.
• Both Forward and Reverse Primers are used in polymerase chain reactions.
• Both Forward and Reverse Primers are synthesized commercially.
• Both Forward and Reverse primers are temperature stable and they are normally having similar Tm.
• Both Forward and Reverse Primers are annealed with the target DNA sequences.
• Both Forward and Reverse Primers are designed according to the PCR reactions.
• Both Forward and Reverse Primers are served as starting points for the DNA amplification.
• Both reverse and forward primers are important for the production of million copies of particular regions of targeted or interested DNA sequences.

What is the Difference Between Forward and Reverse Primer?

Summary – Forward vs Reverse Primer

There are two types of primers involved in PCR technique. They are forward and reverse primers. Based on the elongation of the primer in new DNA strand synthesis, these primers are labeled or named. Taq polymerase synthesizes the new DNA in 5’ to 3’ orientation. Hence, primers are designed as complementary to 3’ ends of the double strands. Forward primer which elongates in 5’ to 3’ direction hybridizes with the 3’ end of the antisense or the template or the noncoding sequence. It serves as the starting point to synthesize coding sequence. Reverse primer which elongates in 5’ to 3’ direction hybridizes with the 3’ end of the coding or the nontemplate or the sense strand. It serves as the starting point to synthesize the noncoding sequence. This is the difference between forward and reverse primer.

Reference:

1.“Primer (Molecular biology).” Wikipedia, Wikimedia Foundation, 20 Feb. 2018. Available here 
2.“Polymerase chain reaction (PCR).” Khan Academy. Available here 

Image Courtesy:

1.’Primers RevComp Melted2’By Richard Wheeler (Zephyris) – Own work, (CC BY-SA 3.0) via Commons Wikimedia