THE REVOLUTION OF Gene Editing TOOLS

Gokul Bhaskaran

BS-MS 5th year, Biological sciences

Gene-editing technology is one of the exhilarating fields that arose a few decades ago and paved the way for a scientific revolution. Artificial selection on plants and animals obtained through random mutations, have become very useful and meaningful. Increase in yield, resistance to disease and the overthrow of certain environmental fluctuations are all made possible by this engineering success.

The skill of selectively targeting the desired genotype/characteristic opened up a new phase of opportunities. The knowledge of DNA structure and function laid the groundwork for this entire field which subsequently witnessed several breakthroughs. At present, gene engineering produces enormous products to study diseases and other therapeutic strategies by efficiently programming cells outside human via a system that mimics the same environment and thus gives an output. The extensive use of prokaryotes make all these manipulations easy and fast. Undoubtedly, gene editing – the way of making changes in specific parts of genome is an art, but the proper incorporation of scientific knowledge is quintessential in achieving faster, cheaper, and more precise methods to add, remove, or change genes in living organisms.

The foundation of gene editing was possible by targeted nucleases with the capability to manipulate any genomic sequence, enabling the easy creation of cell lines and animal models which promoted targeted and effective possibilities for gene therapy. The combination of different zinc finger DNA binding domains that target specific sequences tethered to a FOK1 restriction endonuclease, the zinc finger nuclease (ZFN) was developed as the first programmable nuclease for gene-editing purposes. Very soon ZFN turned into a handy tool to manipulate the genomes of many plants, animals and various types of mammalian cells. But the drawbacks in affinity and assemblage of zinc finger domains made it difficult for non-specialists to routinely engineer ZFNs. The discovery of the DNA binding domains from the plant-bacteria Xanthomonas, brought about the development of the next phase of gene-editing technology.

Transcription activator-like effector nuclease (TALENS) marked another new arrival into the team which can be generated by assembling certain TALE DNA binding domains into the desired combination to target specific sequences. Although designing TALENS was simple, the ease was counteracted by the possibility of lethality due to binding at off-target sites and the induction of undesired DNA cleavage. Both ZFN and TALEN technologies served as the means to edit genes and make targeted changes to DNA in a highly specific and precise manner. But, excessive time consumption in protein engineering and the realization of resource and cost make these gene-editing tools preferable only under extreme requirements.

Overview of TALENS

Source : Wikimedia Commons By Ogletreerd

The latest entry: CRISPR-Cas9 system (clustered regularly interspaced short palindromic repeats- CRISPR-associated protein 9) is a part of microbial immune systems used to defend against foreign DNA, such as from viruses. The non-requirement of protein engineering to direct the nuclease to the target site, using instead just a short piece of RNA to do the editing both in vitro and in vivo began to unravel cellular pathways and deemed this system a powerful and contemporary gene- editing tool.

The CRISPR-Cas9 system can potentially be adopted to replace defective genes that cause disease and this flexibility for gene editing is a new milestone in the genome editing arena. The applications (and controversies) of CRISPR-Cas9 have been quite extensive. It is safe to say that ZFN, TALENS and CRISPR-Cas9 system triggered a revolution and the establishment of a field that changed the face of scientific research.

Different Generations of Gene Editing Tools

Source : Wikimedia Commons By Mazhar Adli - review article : The CRISPR tool kit for genome editing and beyond.

But the whole essence of “how-to”, “where to” and “what for” the gene tools to be used is still a big question to be answered. The plethora of applications and features warrant the further use of editing technology but there is always a dark side. The only way to tackle this is by incorporating the proper management of ethics and scientific knowledge wisely.

Every second brings up changes, let’s anticipate the new changes that will be brought about by gene-editing techniques. All of this will create a fruitful revolution that shapes one of the best advancements for all living beings!

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