What is the Difference Between crRNA tracrRNA and gRNA

The key difference between crRNA tracrRNA and gRNA is that crRNA is one of the two types of RNA of CRISPR, which is complementary to the target DNA sequence, while tracrRNA is the second type of RNA of CRISPR, which serves as a binding scaffold for the Cas nuclease and gRNA is one of the two main components of CRISPR-Cas9 system of bacteria and archaea which recognizes target DNA and guides Cas proteins to make double-strand breaks in target DNA.

CRISPR stands for clustered regular interspersed short palindromic repeats. It is one of the immune systems in bacteria and archaea. Bacteria and archaea use this system to protect themselves from viral pathogens. It is one of the most amazing mechanisms which involves a successful genetic modification. This mechanism (CRISPR-Cas9) is used as a novel gene-editing tool in Biotechnology. It has brought new advances to biotechnology, especially in the field of genetic engineering.

There are two main components of the CRISPR-Cas9 system. They are guide RNA (gRNA) and CRISPR-associated (Cas) nucleases. Guide RNA is a specific RNA sequence that guides Cas proteins to cleave target DNA. Guide RNA consists of two types of RNA. They are crispr RNA (crRNA) and tracrRNA. CRISPR-associated (Cas) nucleases are non-specific endonucleases that make double-strand breaks in target DNA. By making double-strand breaks in target DNA, bacteria and archaea direct their endogenous repair mechanisms to inactivate viral DNA.

CONTENTS

1. Overview and Key Difference
2. What is crRNA  
3. What is tracrRNA 
4. What is gRNA
5. Similarities – crRNA tracrRNA and gRNA
6. crRNA vs tracrRNA vs gRNA in Tabular Form
7. Summary – crRNA vs tracrRNA vs gRNA

What is crRNA?

crRNA or Crispr RNA is one of the two types of guide RNA. Structurally, it is a 17-20 nucleotide sequence. The most valuable feature of crRNA is that it is complementary to the target DNA. Therefore, crRNA matches with the viral DNA sequence. The specificity of the CRISPR-Cas 9 system depends on crRNA. In bacteria, crRNA exists fused to the tracrRNA sequence, which is the second type of CRISPR RNA. crRNA production is triggered by the re-exposure of bacteria to a virus. When exposed, transcription of the gene that is coding for crRNA takes place. Then, the defence mechanism starts against the virus.

What is tracrRNA?

Trans-activating crRNA or tracrRNA is the second part of guideRNA or CRISPR RNA. It is pronounced as tracer RNA.  tracrRNA serves as the binding scaffold for the endonuclease Cas 9 protein. In other words, tracrRNA works as the handle to guide Cas9 towards the target DNA. Structurally, tracrRNA has 42 nucleotides. It exists combined with crRNA.

What is gRNA?

Guide RNA or gRNA is one of the two major components of the CRISPR-Cas9 immune system. It is a specific RNA sequence composed of two elements: crRNA and tracrRNA. gRNA recognizes the target DNA and directs Cas proteins to make double-strand breaks in target DNA. In order to do it, crRNA consists of a complementary sequence of target DNA while tracrRNA guides Cas proteins working as a handle.

Designing the correct gRNA is a critical step in the CRISPR-Cas9 gene-editing tool. Therefore, the success and the editing efficiency of the CRISPR system depend on the correct sequence of gRNA. gRNA can be expressed in cells from a transfected plasmid. When cloned plasmids are introduced into cells, host cells generate gRNA. The most commonly generated gRNA comprises 100 base pairs.

What are the Similarities Between crRNA tracrRNA and gRNA?

• crRNA, tracrRNA and gRNA are RNA sequences found in bacteria and archaea.
• They all belong to the CRISPR system.
• gRNA is made from crRNA and tracrRNA.
• They are involved in recognizing the bacteriophage DNA and guiding endonucleases towards target DNA.
• Hence, all three types of RNA are involved in directing the Cas9 nuclease to make double-strand breaks in invading foreign genetic materials.
• The success of the CRISPR experiment depends on all three types of RNA.

What is the Difference Between crRNA tracrRNA and gRNA?

crRNA is a part of gRNA of CRISPR that is complementary to the target DNA, while tracrRNA is the second part of gRNA, which serves as a binding scaffold for the Cas nuclease. gRNA is the combination of crRNA and tracrRNA, and it is one of the two components of CRISPR-Cas9 system, which recognizes target DNA and guides nucleases to make double-strand breaks in target DNA. So, this is the key difference between crRNA tracrRNA and gRNA.

The following table summarizes the difference between crRNA tracrRNA and gRNA.

Summary – crRNA vs tracrRNA vs gRNA

CRISPR-Cas9 system is an immune system of bacteria and archaea. This system is used as a gene-editing tool in Biotechnology. CRISPR consists of two types of RNA, namely CRISPR RNA (crRNA) and transactivating CRISPR RNA (tracrRNA). They are collectively known as guideRNA or gRNA. This RNA recognizes target sequences of invading pathogens and guides endonucleases to make double-strand breaks in the target DNA. Once double-strand breaks are made in the foreign genetic material, endogenous repair mechanism (nonhomologous end joining; NHEJ) inactivates the foreign DNA by introducing mutations. Thus, this summarizes the difference between crRNA tracrRNA and gRNA.

Reference:

1. “Guide RNA.” An Overview | ScienceDirect Topics.
2. Karvelis, Tautvydas, et al. “CrRNA and Tracrrna Guide cas9-Mediated DNA Interference in Streptococcus Thermophilus.” RNA Biology, Landes Bioscience, May 2013.

Image Courtesy:

1. “CRISPR-Cas9 mode of action” By ViktoriaAnselm – Own work (CC BY-SA 4.0) via Commons Wikimedia
2. “GRNA-Cas9” By Marius Walter – File:GRNA-Cas9.png (CC BY-SA 4.0) via Commons Wikimedia