CRISPER CAS

INTRODUCTION Genome can be edited by removing, adding or

altering sections of the DNA sequence.   There are so many technologies that enable

geneticists and medical researchers to edit parts of the genome. 

These tools allow researchers to permanently modify genes in living cells and organisms and to correct mutations at precise locations in the human genome to treat genetic causes of disease.

CRISPR-Cas9 is a genome editing tool. CRISPR has rapidly become one of the most

popular approaches for genome engineering.

CRISPER CAS

“CRISPR” stands for Clustered Regularly Interspaced Short Palindromic Repeats.

CRISPRs, in conjunction with CRISPR associated (Cas) proteins.

Cas9 is the nuclease guided by the crRNA and tracrRNA (or trans-activating crRNA) to cleave specific DNA sequences.

It is currently the simplest, most versatile and precise method of genetic manipulation.

DISCOVERY

CRISPRs were first discovered in archaea and later in bacteria by Francisco Mojica who proposed that CRISPRs serve as part of the bacterial immune system, defending against invading viruses. The system serves as a genetic memory that helps the cell detect and destroy when they return.

In January 2013, Feng Zhang at the Broad Institute and MIT published the first method to engineer CRISPR to edit the genome in mouse and human cells.

MECHANISM The CRISPR-Cas9 system consists of two key

molecules- 1) An enzyme called Cas9- This acts as a pair of ‘ molecular scissors’ that can cut the two strands of DNA at a specific location in the genome so that bits of DNA can then be added or removed.  2) A piece of RNA called  guide RNA (gRNA)- This consists of a small piece of pre-designed RNA sequence (about 20 bases long) located within a longer RNA scaffold. The scaffold part binds to DNA and the pre-designed sequence ‘guides’ Cas9 to the right part of the genome. This makes sure that the Cas9 enzyme cuts at the right point in the genome.

In DNA there is a specific targeted part.

Steps

The guide RNA will bind to the target sequence and no other regions of the genome. Actually the guide RNA is designed to find and bind to a specific sequence in the DNA.

The Cas9 follows the guide RNA to the same location in the DNA sequence and makes a cut across both strands of the DNA.

At this stage the cell recognises that the DNA is damaged and tries to repair it. The DNA repair machinery is used to introduce changes to one or more genes in the genome of a cell of interest.

Steps

Several ‘gene editing’ technologies have recently been developed to improve gene targeting methods, including CRISPR-Cas systems, transcription activator-like effector nucleases (TALENs) and zinc-finger nucleases (ZFNs). 

The CRISPR-Cas9 system currently stands out as the fastest, cheapest and most reliable system for ‘editing’ genes.  

Other Techniques For Gene Altering

It is used as a tool for treating a range of medical conditions that have a genetic component, including cancer, hepatitis B or even high cholesterol. 

Many of the proposed applications involve editing the genomes of somatic (non-reproductive) cells but there has been a lot of interest in and debate about the potential to edit germline

(reproductive) cells.

Because any changes made in germline cells will be passed on from generation to generation it has important ethical implications. 

Wang et al. (2014) used both TALEN and CRISPR/Cas9 technologies to target and successfully knock out the genes of the mildew-resistance locus (MLO) in wheat to generate plants resistant to powdery mildew disease.

Applications

Undoubtedly this process caught most attention for their potential in medical applications and numerous other biotechnological applications like crop editing, gene drives and synthetic biology

Despite the enormous potential that lies within the CRISPR-Cas9 technology, further investigation is required to make the system an applicable and safe tool for therapeutically useful approaches. 

Conclusion