Research led by Wardah Shahzadi

Shahzadi Fatima, Aamna Asim, Fatima Shamim

Introduction:

CRISPR (Clustered Regularly Inter-Spaced Short Palindromic Repeats”) is a groundbreaking tool that has revolutionized genetics. Going further we will learn more about it and its relationship with Genetic Engineering.

 Discovery of CRISPR:

CRISPRs were first discovered in Archaea (and later in bacteria) by Francisco Mojica, a scientist at the University of Alicante in Spain. He proposed that CRISPRs serve as part of the bacterial immune system, defending against invading viruses. They consist of repeating sequences of genetic code, interrupted by “spacer” sequences (crucial for the system’s function) – remnants of genetic code from past invaders. The system serves as a genetic memory that helps the cell detect and destroy invaders (called “Bacteriophage”) when they return. This discovery has opened pathways to various applications beyond its original purpose.

 How does CRISPRs work?

Scientists realized they could use CRISPRs to edit DNA with incredible accuracy. Imagine having a pair of molecular scissors, guided by a custom-made GPS, that can cut exactly where you want in a gene. That’s essentially how CRISPR works but here's a better outline:

CRISPR “spacer” sequences are transcribed into short RNA sequences (“CRISPR RNAs” or “crRNAs”) and this guides the system to specific DNA sequences for editing (matching sequences of DNA). When the target DNA is found, Cas9 – one of the enzymes produced by the CRISPR system –binds to the DNA and cuts it, shutting the targeted gene off. Using modified versions of Cas9, researchers can activate gene expression instead of cutting the DNA. These techniques allow researchers to study the gene’s function. In simpler terms, your DNA can get a customization of its own.

Advantages of CRISPRs in Genetic Engineering:

1)         CRISPR technology enables for gene editing/ genetic modifications.

2)        Genetically engineered crops require less fertilizers and water, making the farming process more Eco-Friendly.

3)        Some types of Genetically engineered crops are even insect resistant eliminating the need for insecticides that contain harmful chemicals. Countries like Pakistan that are mainly agriculture based and have to import their fertilizers would benefit immensely from crops that do not require fertilizer or a very little amount.

4)        CRISPR as a tool is fast efficient and flexible.

5)        CRISPR-Cas9 can be used to target and modify “typos” (aka mutations) in the three-billion-letter sequence of the human genome in an effort to treat genetic disease.

6)        CRISPR-Cas9can edit multiple genes simultaneously, when the implemented sgRNAs are designed to target different genetic loci, which makes it perfect for manipulating multiple locations in the genome. The simplicity of CRISPR-Cas9 –relying largely on 2 molecules to induce edits – as well as the flexibility available in designing sgRNAs to bind to any/numerous target(s) means that researchers can start editing quickly and make adjustments easily. Which is beneficial especially during the trouble shooting phases of research enabling meaningful advancements at an accelerated pace.

 Disadvantages of CRISPRs:

1)         There is risk of contaminating ecosystems in regard to genetically engineered organisms spreading and reproducing in the wild which could alter ecosystems affecting biodiversity in extreme unpredictable ways.

2)        It is generally believed that addition of genetic material as in transgenic crops the foreign DNA has contaminated the crop and is unsafe for human use, this however is not backed by scientific evidence.

3)        CRISPR itself has its limitations CRISPR-Cas9has to be successfully delivered into cells in order to induce the desired edits. Delivery of the Cas9/gRNA complex into a sufficient number of cells can be challenging because all of the components must be delivered into cells at the right concentrations and at the correct point in a cell's cycle this creates a sizable room for error.

4)        It has efficiency limitations as the gene editing activity itself may not occur once even if the editing complex is taken up by the cell, especially when inserting material into a gene.

5)        Off-target effects occur when the Cas9nuclease edits an untargeted section of the genome, resulting in unwanted alterations. These effects are a significant concern in CRISPR-Cas9 experiments and they can be challenging to predict.

 Ethical Concerns:

CRISPR/Cas9 has shown incredible potential despite the fact that it is less than a decade old however, there has been obvious bio-ethical concerns regarding the use of it. A major controversy linked to CRISPR technology is the possible application of it on human embryos and it is because till this day there is a lack of clarification on the status of the human embryo. Different scientific communities around the globe view the time period of when to start considering an embryo a person differently since religious groups, governments, funding agencies or just a panel of experts will have different opinions leading to cloudy judgement which could result in a public outcry. Off target mutations is another very controversial concern since it is possible for it to have canceling effects on humans as well as the environment. Larger genomes can have multiple identical sites and hence CRISPR can cut sequences that are not calculated. Such it is that it could lead to cell death as well as abnormal transformations. Scientists have put warnings regarding the risks of accidental release in the environment of experiment organisms modified using gene drive. Regarding regulations of the modified organisms, it is difficult to identify the respective modified organisms outside the laboratories. CRISPR gene editing in the germ lines is banned around the world due to safety reasons (except in the UK since February 2016).

 Future Potential:

CRISPR genome editing has vividly allowed scientists and researchers around the world to be two steps ahead of diseases like cancer and mental illnesses as they now acquire the ability to rush the creation of animal and cell models. CRISPR is being used as a diagnostic and in treatment of new and unknown pathogens. It allows scientists to carry out larger and more complex experiments that are vital to understand features of pathogens. A successful example of CRISPR modification is sickle cell anemia. The existing treatment is bone marrow transfusions from a healthy person but since this disease is genetic CRISPR has given hope for new treatments that prevent the introduction of foreign bodies in the recipient that the body may reject or the donor cells attacking the recipient. All in all, CRISPR is a promising and powerful tool that can and will show wonders in the near future that will make vast improvement in the world of health sciences and medicine. It's so precise, cheap, and easy to use such that CRISPR is already making waves in medicine, helping to develop treatments for genetic disorders, and in agriculture, creating more resilient crops. But with its incredible potential comes a lot of ethical questions, especially when it comes to editing human genes. As exciting as CRISPR is, it also makes us think deeply about how we should use this power to shape life itself.

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