Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated protein 9. A basic CRISPR-Cas9 system consists of a Cas9 endonuclease and a small RNA that guides Cas9 to the target DNA.

Origin

CRISPR sequences were first observed in bacteria and later identified in archaea. Researchers discovered that the CRISPR-Cas9 system serves an adaptive immune defense against invading viruses. Many bacteria and most archaea capture short sequences of the viral DNA to create a library of virus DNA segments, or CRISPR arrays. When the prokaryotes are re-exposed to the same virus or class of viruses, CRISPR arrays are used to transcribe small RNA segments that help recognize viral invaders and subsequently destroy viral DNA with Cas9 or a similar endonuclease.

Using CRISPR-Cas9 Technology

CRISPR-Cas9 is commonly used in the laboratory to remove DNA and insert a new DNA sequence in its place. To achieve this, researchers must first create a small fragment of RNA called the guide RNA, with a short sequence called the guide sequence that binds to a specific target sequence on genomic DNA. The guide RNA can also associate with Cas9 (or other endonucleases like Cpf1). The guide RNA and Cas9 protein are administered to a cell of interest where the guide RNA identifies the target DNA sequence and Cas9 cleaves it.

The cell’s machinery then repairs the broken strands by inserting or deleting random nucleotides, rendering the target gene inactive. Alternatively, a customized DNA sequence may be introduced into the cell along with the guide RNA and Cas9, that serves as a template for the repair machinery and replaces the excised sequence. This is a highly effective way for researchers to “knock out” a gene to study its effects or replace a mutated gene with a normal copy in hopes of curing a disease.

Ethical and Feasibility Considerations in Humans

As a result of the significant gene modification capabilities of the CRISPR-Cas9 system, there has been great debate over its use, especially in regards to embryo editing. A Chinese scientist recently claimed to have created genome-edited babies using CRISPR technology to disable a gene involved in HIV infection. This led to a global outcry from scientists concerned about the ethical and safety considerations of the procedure. Many have called the move premature, and others have expressed concerns over off-target genomic effects. While the number of possible biotech applications for the CRISPR-Cas9 system is numerous, it is important to consider future challenges that may arise as a result of its use.