“Genome editing” the word itself carries so much fascination within itself. Think about the wonders it does. Genome editing has vast, limitless applications in today’s world. It is making miracles happen on day to day basis. Well, a question comes to our mind, What makes genome editing so simple, versatile and yet so precise? Among all the other technologies brewing on the surface like Zinc Finger Nuclease(ZFNs), TALENs the technology which is responsible for changing the face of genome engineering with much ease is CRISPR Cas9 technology. The advent of this miraculous tool CRISPR Cas9 technology has ignited a new revolution in the world of genome engineering.
- CRISPR stands acronym for Clustered Regularly Interspaced Short Palindromic Repeats. Cas 9 is CRISPR- associated protein 9.
- CRISPR palindromic repeats along with Cas9 nuclease play an important role in microbial immunity.
- CRISPR is found in approximately 50% of sequenced bacterial genomes and nearly 90% of the sequenced archaebacteria.
- These sequences are derived from the fragments of bacteriophages that had previously infected the prokaryote.
- They are used to detect and destroy DNA from similar bacteriophages during subsequent infections.
- Hence the sequence plays a key role in the antiviral (i.e, Antiphage) defence system of the prokaryotes and provide a form of acquired immunity.
What is Cas9?
- Cas 9 is an enzyme that uses CRISPR sequences as a guide to recognize and cleave specific strands of DNA that are complementary to CRISPR sequences.
- Many variants exist with different functions (SSB, DSB, etc) due to Cas9’s DNA recognition function.
- The protein typically binds to two RNA molecules: cr RNA and another called tracrRNA( or “ trans-activating crRNA”).
- Analogous to the search function in the modern word processors, Cas9 can be guided to specific locations within complex genomes by a short RNA search string.
- Using this system, DNA sequences within the endogenous genome and their functional outputs are now easily edited or modulated in virtually any organism of choice.
- Cas 9 mediated genetic perturbation is simple and scalable, empowering researchers to elucidate the functional organization of the genome at the systems level and establish casual linkages between genetic variations and biological phenotypes.
CRISPR RNA (crRNA):
Once a spacer is incorporated and the virus attacks again, a portion of the CRISPR is transcribed and processed into CRISPR RNA, or “cr RNA”. The nucleotide sequence of the CRISPR is transcribed and processed into CRISPR acts as a template to produce a complementary sequence of single-stranded RNA. Each crRNA consists of a nucleotide repeat and a spacer portion, according to a 2014 review by Jennifer Doudna and Emmanuella Charpentier, published in the science journal.
WORKING OF CRISPR CAS 9
The following steps elucidate the working of CRISPR CAS 9 in short:-
- 1) Scientists start with RNA. It works as a guide to Cas9. It detects the DNA where editing needs to be done.
- 2) RNA binds to the specific target genome of the DNA. It also binds to the Cas9 enzyme. The Cas9 then locks up onto the double-stranded DNA and unzips it.
- 3) Once the DNA is cut, researchers use the cell’s DNA repair machinery to add or delete pieces of genetic material, or to make changes to the DNA by replacing an existent segment with a customized DNA sequence.
USES OF CRISPR CAS 9 TECHNOLOGY IN VARIOUS FIELDS:
- THE STATUS OF PIG PRODUCTION AND CURRENT APPLICATION OF CRISPR Cas9 TECHNOLOGY:
Worldwide pig(Su’s scrogs domestics) production accounted for 42% of the total livestock production in 2018 and this percentage is expected to go up by 2050. Pork is consumed by a large population around the world.
Using the CRISPR Cas9 technology is benefitting the quality and quantity of pork production.
In addition to primates, it has been found that pigs share similar characteristics with humans such as organ size, genome length, blood glucose levels, complexity and composition of chromosomes.
So, not only pigs are of great domestic use but also can be used in the labs for testing food and pharmaceutical applications.
- CRISPR technology is also used in the editing of stem cells.
- Researchers are trying to find a cure for cancer, blindness, cardiovascular diseases, Alzheimer’s disease, HIV, Haemophilia, cystic fibrosis, sickle cell anaemia and many more using this technology.
- IN PLANT BIOTECHNOLOGY
- DESIGNER BABY
WHAT ARE THE ETHICAL CONCERNS?
Ethical issues arise when this technology is used to edit genomes in germline cells. As long as genome alterations are concerned to stem cells in which the changes are not passed from one generation to other. So Germ line cells and embryo editing bring up ethical challenges.
Throughout the article, it has been stressed enough about the uses of CRISPR Cas9 technology and its pros. It can be said that with the invention of this technology many problems have been solved with ease. The only cons stand in the way that it should not be used in any unethical way. Since it is a very powerful biological weapon.