Neha BaruahIn a remarkable leap forward, scientists at the CSIR-Institute of Genomics and Integrative Biology (IGIB) in New Delhi have pioneered an advanced genome-editing system. This innovative technology surpasses the capabilities and accuracy of the existing CRISPR techniques, promising more precise genetic modifications with fewer unintended consequences.
Understanding CRISPR: The Basics
CRISPR, which stands for Clustered Regularly Interspaced Short Palindromic Repeats, is essentially a bacterial immune system that targets and dismantles viral DNA. Researchers ingeniously adapted this natural mechanism for gene editing in more complex organisms, making it a powerful tool in genetics.
The CRISPR-Cas9 System: How It Works
The CRISPR-Cas9 system has revolutionized genetic research by allowing scientists to add, delete, or modify DNA sequences with a fair degree of accuracy. It uses a guide RNA (gRNA) to lead the Cas9 enzyme to a specific DNA location, where the enzyme cuts the DNA strand. The cell then attempts to repair the break, enabling the introduction of new genetic material or the correction of existing sequences. Despite its revolutionary impact, the CRISPR-Cas9 system can sometimes create “off-target” effects, where unintended parts of the genome are edited, thus posing a challenge for precise genetic engineering.
Transitioning to FnCas9: A Superior Solution
To address these off-target effects, researchers turned their attention to the FnCas9 enzyme derived from the bacterium Francisella novicida. While FnCas9 has shown greater precision and efficiency compared to the commonly used SpCas9, it has historically been less effective overall. The team at CSIR-IGIB enhanced FnCas9 by altering its amino acid interactions with the PAM (Protospacer Adjacent Motif) sequence, thereby boosting its DNA binding capacity and editing efficiency. These modifications have enabled the enzyme to reach more challenging areas of the genome, improving the overall effectiveness of gene editing.
Enhanced Diagnostic Capabilities
The improved FnCas9 enzyme demonstrated remarkable capabilities in identifying specific single-nucleotide changes in DNA, effectively doubling the detection of genetic variations linked to diseases. Testing on human kidney and eye cells revealed that the enhanced FnCas9 exhibited higher precision and significantly fewer off-target effects compared to the SpCas9 enzyme. This suggests significant potential for treating a variety of genetic disorders with greater accuracy and safety.
Case Study: Leber Congenital Amaurosis 2 (LCA2)
A striking application of the enhanced FnCas9 was in correcting a mutation in the RPE65 gene, which causes Leber Congenital Amaurosis 2 (LCA2), a severe form of childhood blindness. The new system almost completely corrected the genetic mutation, resulting in normal protein production in retinal cells. This success was notably more effective than previous CRISPR systems. Building on these promising results, scientists are exploring the use of patient-derived stem cells. By editing these stem cells to correct mutations before transplanting them back into the patient, this method could provide a safer and more precise alternative to direct CRISPR application.
Neha Baruah
