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Research blast on gene editing

Gene editing is a relatively new biological technique that has the potential to treat everyone with cystic fibrosis (CF) in the future. It works by altering the DNA sequence of the CFTR gene to correct mutations within it. In this issue of Research Blast – your guide with a difference to CF research – we delve into some of the details of this technique.

Q What links gene editing with the Olympics?

A The Summer of 2012!

At the end of June 2012, a month before the iconic opening ceremony of the 2012 London Olympics, a research paper was published online by Professors Jennifer Doudna and Emmanuelle Charpentier and their colleagues that would revolutionise gene editing research.

doodle of gene editing

They used a biological technique, adapted from a bacterial defence system, to cut DNA in a very specific place: a technique called CRISPR-Cas9, which is now used extensively in gene editing research. The ‘Cas9’ part of this technique are the ‘scissors’ that make the cut in the DNA, and the ‘CRISPR’ part is a way of guiding the scissors to the right place.

Once the cut is made, the repair can be carried out, whether it is removing bits of DNA that shouldn’t be there or replacing a damaged section with a new piece of undamaged DNA. The cuts are repaired by normal processes we already have in our body. It’s vital that the only part of DNA that is cut is the part you’re trying to edit – cuts in DNA elsewhere could cause unwanted side effects.

 

Q What does gene editing share with the legs of a caterpillar?

A The number 16

An important part of the CRISPR-Cas9 gene editing system is that it can be adapted to cut a specific piece of DNA. The way to do this is to design a ‘guide’ template that will direct the scissors to the right place. The guides are made from building blocks of RNA, which pair up with the building blocks of DNA. The first rule of designing a ‘guide’ RNA is ensuring it is long enough to be unique.

Researchers have calculated that it needs to be at the very least 16 building blocks long. Any shorter, and there’s a chance another section of DNA with a similar combination of building blocks could be mistakenly cut. (In reality, 20 building blocks are used to make the guides, allowing for an extra bit of ‘back up’). If you want to know how the maths is worked out… read on!

There are 3 billion (that’s nine zeros) building blocks in the length of our entire DNA. If the ‘guide’ RNA was four building blocks long, there would be 256 ways these blocks could be arranged. However, in the 3 billion building blocks of our DNA, there would be 11 million other places (i.e. 3 billion divided by 256) with the same four block code. This means the DNA will be cut in over 11 million ways – it would be cut to shreds!

Even increasing the length of the CRISPR-Cas9 guide to 12 building blocks would still be too short, as the DNA could be cut in nearly 180 different places. It is only by making it at least 16 building blocks long that researchers can be confident enough to know the guide will only cut where they want it to – i.e. the CFTR gene.

 

Q Why is gene editing like getting a new sofa?

A You have to think about the delivery!

One of the questions that researchers in our gene editing Strategic Research Centre (SRC) have been investigating is how to get the gene editing machinery into the lung cells in order to correct the mutated CFTR gene. The machinery that needs to be delivered includes the ‘guide’ RNA; the molecular ‘scissors’ to cut a hole in the DNA to make the repair; and the new section of DNA to be added to the gene.

The research team have tested whether gene editing works better if a larger, fully assembled version of the gene editing machinery is delivered to the site of the repair, or whether it is possible to develop and deliver the molecular equivalent of flatpack furniture, which would be assembled on site within the target cells. The ‘flatpack’ is smaller to package up, which is a benefit, but the research team wanted to find out if the complexity of assembling the machinery ‘onsite’ would outweigh this advantage. The researchers have found that, while both systems worked well, the fully assembled approach may produce less side effects.

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