ABC Science By Bernie Hobbs Updated 11 Apr 2016, 11:03pm


A powerful new gene-editing technology called CRISPR has enormous potential to treat human diseases but the ability to tinker with genes can also be controversial. Here we explain what CRISPR is and how it works.

Since gene technology first emerged over 40 years ago we’ve seen a wealth of genetic advances — not least of all the decoding of the human genome in 2001.

Key points:

  • CRISPR is a faster, cheaper and more accurate way of editing genes
  • It should allow replacing faulty genes with healthy ones
  • CRISPR is not yet accurate enough to use on people
  • Scientists have called for a moratorium on using CRISPR on sperm and egg cells

But that’s nothing compared to the genetic revolution that we’re at the beginning of right now, thanks to a technique adapted from bacteria called CRISPR (the catchy acronym for clustered regularly interspaced short palindromic repeats).

Researchers learn what genes do by switching them on or off, or cutting them out of the DNA in a cell entirely.

Since it appeared in 2012, CRISPR has completely transformed the process that researchers use to edit genes this way.

It’s not the first method devised for this kind of genome editing, but CRISPR is a lot cheaper, faster, and more accurate than any of the alternatives. In technology jargon, it’s a capital D disruptor.

And with applications in gene therapy (replacing faulty genes with healthy ones), drug research and agriculture for starters it’s no wonder the method has taken off like a rocket.

What is CRISPR? And how does it work?

Editing genes can mean removing or replacing an existing gene, switching a gene on or off, or inserting a new gene altogether.

Whatever the aim, the first step is always to find the stretch of DNA that codes for the gene and grab hold of it, so a cut or tweak can be made.

CRISPR not only finds the target gene and locks on, it also delivers an enzyme that cuts the DNA. And it does all this with unprecedented accuracy.

The reason it’s able to manage this precision double act is because CRISPR is made of ribonucleic acid (RNA) — a molecule that can be tailor-made to perfectly match a sequence of DNA or to bind to a protein.

CRISPR RNA does both jobs — one end is custom-made to match the target gene’s DNA sequence, and the other end binds to a DNA-cutting enzyme, or nuclease.

CRISPR-Cas9 attacks DNA
INFOGRAPHIC: CRISPR comprises a single molecule of RNA (shown here in purple) that performs two jobs. One end binds to the target gene (dark red), the other end delivers a DNA-cutting enzyme (in this case Cas9) to the site.

 It’s a brilliant system, and it wasn’t cooked up in a lab — scientists pinched it from bacteria.

Simple beings that they are, bacteria have a version of an immune system, and CRISPR is at the heart of it.

When a virus invades a bacterial cell, it leaves traces of its DNA in the bacterial genome. If the bacterium encounters that virus again, CRISPR RNA uses the viral DNA remnants and a nuclease called Cas9 to attack the virus.

An improved version of this CRISPR-Cas9 combination is now being used in laboratories around the world.

Cas9 is not the only nuclease in the game — there are a number of Cas (CRISPR-associated) proteins, each with a slightly different capability.

Researchers simply order the sequence of guide RNA to include a part of the gene they’re interested in, plus the Cas binding sequence, mix it with the Cas protein to suit the job and they’re ready to go.

What can CRISPR do that other gene technologies can’t?

CRISPR has two main rivals in the genome editing game, with equally odd names: zinc-finger nucleases and TALENS.

Like CRISPR, these gene editing systems can deliver a DNA-cutting enzyme to a particular gene.

But they have one key difference: while CRISPR is made of an RNA molecule, zinc-finger nucleases and TALENS are proteins, which are much trickier molecules to work with.

Unlike RNA, it’s not just the sequence of a protein that matters when it comes to making it bind — the shape is critical.

If a protein isn’t just the right shape with positive and negative charges in just the right positions, it won’t stick to DNA.

So tailoring a zinc-finger protein or TALE (TAL Effector) protein so that it’s the right shape and charge to bind to a particular gene is far more complex than ordering a custom sequence of RNA that’s a perfect match.

And complex translates to more time consuming, more expensive and less accurate to work with than CRISPR.

CRISPR technology has another benefit — it can target multiple genes in a cell at once.

Like in the bacterial system where it originated, multiple CRISPR-Cas9 combinations can exist in the one cell, all targeting different stretches of DNA — it’s just a matter of using different guide RNA sequences. That “multiplexing” just isn’t possible with protein-based guide systems.

Considering that most diseases and conditions involve more than a single gene, multiplexing opens the way to studying and potentially treating more complex genetic issues down the track.

What are the limitations of CRISPR?

While it’s a game-changer in gene editing technology, CRISPR-Cas9 isn’t error proof.

It has a much higher success rate than the other nuclease technologies when it comes to cutting DNA at the right place. But like the other systems it can also make unintended cuts outside the target gene.

It’s still early days in the technology, and reducing these off-target errors is a big focus for those working on improving CRISPR. A slight change in the Cas9 protein has already shown a significant reduction in off-target errors — the technique will need to be incredibly robust before it’s applied in a clinical setting.

Where is CRISPR at now?

CRISPR-Cas9 is still very much confined to research laboratories, but things are changing so rapidly in this field that this section will need regular updating.

To date, the system has been used to edit genes in every one of the standard model organisms (like fruit flies, zebrafish, frogs and mice) that researchers generally work with.

It’s also been used in mice to successfully disable the gene that causes Huntington’s disease, and to prevent muscular dystrophy. CRISPR has also allowed deletion of 60 viruses from pig genes, paving the way for lower risk transplant of pig organs into humans.

Plans are afoot to use CRISPR to delete the gene for the protein that the HIV virus uses to enter T cells in the immune system, effectively locking the virus out.

Deleting genes is one thing, but inserting replacement genes is a taller order.

Success rates for this step have only just started nudging 60 per cent — well short of the extreme accuracy that is required for any clinical applications, like replacing a faulty disease-causing gene with a correct version, or engineering a cell so that it’s impervious to a virus like HIV.

Why is CRISPR controversial?

As with all methods that let us directly change genes, CRISPR has raised alarm bells on a few fronts. And its rapid uptake across the board in biotech research has some scientists understandably concerned that we’re racing ahead with experiments before knowing the full implications of the technology.

Nowhere is this more evident than in work involving human embryos.

The announcement in 2015 that Chinese scientists had edited the genome of an embryo drew global attention to the issue. (The embryo was un-viable — physically incapable of developing into a fetus or human.)

In 2016 approval was also given to a British biologist to use CRISPR on unwanted human embryos to better understand the role of genes in healthy development. (The researchers will use unwanted IVF embryos and the experiments — and embryos — will be terminated after one week.)

While the embryos from these experiments won’t result in a child, they have added urgency to the debate around what limitations need to be put on the use of CRISPR.

That was the focus of the International Summit on Human Gene Editing in Washington in December 2016, which resulted in a call for a moratorium on using CRISPR on germ line cells (egg and sperm) until all safety issues and societal concerns have been addressed. This call has since been backed by a second group of researchers who attempted to edit human embryos.

As the accuracy and safety of CRISPR improves, and the potential clinical benefits of CRISPR become feasible, this stance will certainly evolve.

Bookmark and Share